Phenotypic, transcriptomic and genomic characterization of clonal plasma cells in light chain amyloidosis [Gene expression profiling]
Ontology highlight
ABSTRACT: Immunoglobulin light-chain amyloidosis (AL) is a rare clonal plasma cell (PC) disorder that remains largely incurable. AL and multiple myeloma (MM) share the same cellular origin, but while knowledge about MM PC biology has improved significantly, the same does not apply for AL. Here, we undertook an integrative phenotypic, molecular, and genomic approach to study clonal PCs from 22 newly-diagnosed AL patients. Through principal-component-analysis, we demonstrated highly overlapping phenotypic profiles between AL and MGUS or MM patients. However, in contrast to MM, highly-purified FACSs-sorted clonal PCs in AL (n=9/22) show virtually normal transcriptomes with only 68 deregulated genes as compared to normal PCs, including a few tumor suppressor (CDH1, RCAN) and pro-apoptotic (GLIPR1, FAS) genes. Notwithstanding, clonal PCs in AL (n=11/22) were genomically unstable with a median of 9 copy-number-abnormities (CNAs) per case; many of which similar to those found in MM. Whole-exome sequencing (WES) was performed in three AL patients and revealed a median of 10 non-recurrent mutations per case. Altogether, we showed that although clonal PCs in AL display phenotypic and CNA profiles similar to MM, their transcriptome is remarkably similar to that of normal PCs. First-ever WES revealed the lack of a unifying mutation in AL
Project description:Immunoglobulin light-chain amyloidosis (AL) is a rare clonal plasma cell (PC) disorder that remains largely incurable. AL and multiple myeloma (MM) share the same cellular origin, but while knowledge about MM PC biology has improved significantly, the same does not apply for AL. Here, we undertook an integrative phenotypic, molecular, and genomic approach to study clonal PCs from 22 newly-diagnosed AL patients. Through principal-component-analysis, we demonstrated highly overlapping phenotypic profiles between AL and MGUS or MM patients. However, in contrast to MM, highly-purified FACSs-sorted clonal PCs in AL (n=9/22) show virtually normal transcriptomes with only 68 deregulated genes as compared to normal PCs, including a few tumor suppressor (CDH1, RCAN) and pro-apoptotic (GLIPR1, FAS) genes. Notwithstanding, clonal PCs in AL (n=11/22) were genomically unstable with a median of 9 copy-number-abnormities (CNAs) per case; many of which similar to those found in MM. Whole-exome sequencing (WES) was performed in three AL patients and revealed a median of 10 non-recurrent mutations per case. Altogether, we showed that although clonal PCs in AL display phenotypic and CNA profiles similar to MM, their transcriptome is remarkably similar to that of normal PCs. First-ever WES revealed the lack of a unifying mutation in AL
Project description:Immunoglobulin light-chain amyloidosis (AL) is a rare clonal plasma cell (PC) disorder that remains largely incurable. AL and multiple myeloma (MM) share the same cellular origin, but while knowledge about MM PC biology has improved significantly, the same does not apply for AL. Here, we undertook an integrative phenotypic, molecular, and genomic approach to study clonal PCs from 22 newly-diagnosed AL patients. Through principal-component-analysis, we demonstrated highly overlapping phenotypic profiles between AL and MGUS or MM patients. However, in contrast to MM, highly-purified FACSs-sorted clonal PCs in AL (n=9/22) show virtually normal transcriptomes with only 68 deregulated genes as compared to normal PCs, including a few tumor suppressor (CDH1, RCAN) and pro-apoptotic (GLIPR1, FAS) genes. Notwithstanding, clonal PCs in AL (n=11/22) were genomically unstable with a median of 9 copy-number-abnormities (CNAs) per case; many of which similar to those found in MM. Whole-exome sequencing (WES) was performed in three AL patients and revealed a median of 10 non-recurrent mutations per case. Altogether, we showed that although clonal PCs in AL display phenotypic and CNA profiles similar to MM, their transcriptome is remarkably similar to that of normal PCs. First-ever WES revealed the lack of a unifying mutation in AL A total of 22 patients with confirmed diagnosis of AL based on the presence of amyloid-related systemic syndrome, positive amyloid tissue staining with Congo red, and evidence of PC clonality were studied. Samples were collected after informed consent was given, in accordance with local ethical committee guidelines and the Helsinki Declaration. GEP was performed in 9/22 AL cases with adequate RNA extracted from FACS-purified clonal PCs according to patient-specific aberrant phenotypes, and compared to that of normal PCs from 5 healthy individuals (FACSAriaIIb, BDB; â¥95% purity). RNA was hybridized to the Human Gene 1.0 ST Array (Affymetrix, Santa Clara, CA, USA) and normalization was carried using the expression console (Affymetrix) with the RMA algorithm which includes background correction, normalization and calculation of expression values (log2). Differentially expressed genes between classes were identified using the Significant Analysis of Microarrays (SAM) algorithm (http://www-stat.standford.edu/-tibs/SAM), and significant genes were selected based on the lowest q-value (<10-5).
Project description:Immunoglobulin light-chain amyloidosis (AL) is a rare clonal plasma cell (PC) disorder that remains largely incurable. AL and multiple myeloma (MM) share the same cellular origin, but while knowledge about MM PC biology has improved significantly, the same does not apply for AL. Here, we undertook an integrative phenotypic, molecular, and genomic approach to study clonal PCs from 22 newly-diagnosed AL patients. Through principal-component-analysis, we demonstrated highly overlapping phenotypic profiles between AL and MGUS or MM patients. However, in contrast to MM, highly-purified FACSs-sorted clonal PCs in AL (n=9/22) show virtually normal transcriptomes with only 68 deregulated genes as compared to normal PCs, including a few tumor suppressor (CDH1, RCAN) and pro-apoptotic (GLIPR1, FAS) genes. Notwithstanding, clonal PCs in AL (n=11/22) were genomically unstable with a median of 9 copy-number-abnormities (CNAs) per case; many of which similar to those found in MM. Whole-exome sequencing (WES) was performed in three AL patients and revealed a median of 10 non-recurrent mutations per case. Altogether, we showed that although clonal PCs in AL display phenotypic and CNA profiles similar to MM, their transcriptome is remarkably similar to that of normal PCs. First-ever WES revealed the lack of a unifying mutation in AL A total of 22 patients with confirmed diagnosis of AL based on the presence of amyloid-related systemic syndrome, positive amyloid tissue staining with Congo red, and evidence of PC clonality were studied. Samples were collected after informed consent was given, in accordance with local ethical committee guidelines and the Helsinki Declaration. Genome-wide detection of CNAs and copy-number neutral loss-of-heterozygosity (LOH) were investigated using the Cytoscan 750K platform (Affymetrix) in 11/22 cases with adequate DNA extracted from FACS-sorted clonal PCs and paired T-lymphocytes. The AGCC and ChAS software programs (Affymetrix) were used for data analysis as described elsewhere. CNAs were reported when the three following criteria were met: â¥25 consecutive imbalanced markers per segment; â¥100Kb minimum genomic size and; <50% overlap with paired control DNA and/or genomic variants of Toronto DB (DGV). Only CNN-LOH larger that 5Mb, with â¥25 consecutive imbalanced markers per segment, and <50% overlap with patient-paired CNAs were considered.
Project description:Transcriptional atlas of normal PC development in secondary lymphoid organs (SLO), peripheral blood (PB) and BM for comparison with the transcriptional programs (TPs) of tumor PCs in AL, MM and MGUS based on bulk and single-cell RNAseq
Project description:Transcriptional atlas of normal PC development in secondary lymphoid organs (SLO), peripheral blood (PB) and BM for comparison with the transcriptional programs (TPs) of tumor PCs in AL, MM and MGUS based on bulk and single-cell RNAseq
Project description:Persistence of chemoresistant minimal residual disease (MRD) plasma cells (PCs) relates to inferior survival in multiple myeloma (MM). MRD PCs are therefore a minor clone able to recapitulate the initial tumor burden at relapse and accordingly, its characterization may represent a unique model to understand chemoresistance; unfortunately, the MRD clone has never been biologically investigated. Here, we compared the antigenic profile of MRD vs. diagnostic clonal PCs in 40 elderly MM patients enrolled in the GEM2010MAS65 study, and showed that the MRD clone is enriched by cells over-expressing integrins (CD11a/CD11c/CD29/CD49d/CD49e), chemokine receptors (CXCR4) and adhesion molecules (CD44/CD54). Genetic profiling of MRD vs. diagnostic PCs showed identical copy number alterations (CNAs) in 3/8 cases, 2 patients with linear acquisition of additional CNAs in MRD clonal PCs, and 3 cases with variable acquisition and loss of CNAs over time. The MRD clone showed significant downregulation of genes particularly related to protein processing in endoplasmic reticulum, as well as novel deregulated genes such as ALCAM that is prognostically relevant in MM and identifies chemoresistant PCs in vitro. Together, we show that therapy-induced clonal selection is already present at the MRD stage, in which chemoresistant PCs show a specific phenotypic signature that may result from the persistence of clones with different genetic and gene expression profiles.
Project description:Persistence of chemoresistant minimal residual disease (MRD) plasma cells (PCs) relates to inferior survival in multiple myeloma (MM). MRD PCs are therefore a minor clone able to recapitulate the initial tumor burden at relapse and accordingly, its characterization may represent a unique model to understand chemoresistance; unfortunately, the MRD clone has never been biologically investigated. Here, we compared the antigenic profile of MRD vs. diagnostic clonal PCs in 40 elderly MM patients enrolled in the GEM2010MAS65 study, and showed that the MRD clone is enriched by cells over-expressing integrins (CD11a/CD11c/CD29/CD49d/CD49e), chemokine receptors (CXCR4) and adhesion molecules (CD44/CD54). Genetic profiling of MRD vs. diagnostic PCs showed identical copy number alterations (CNAs) in 3/8 cases, 2 patients with linear acquisition of additional CNAs in MRD clonal PCs, and 3 cases with variable acquisition and loss of CNAs over time. The MRD clone showed significant downregulation of genes particularly related to protein processing in endoplasmic reticulum, as well as novel deregulated genes such as ALCAM that is prognostically relevant in MM and identifies chemoresistant PCs in vitro. Together, we show that therapy-induced clonal selection is already present at the MRD stage, in which chemoresistant PCs show a specific phenotypic signature that may result from the persistence of clones with different genetic and gene expression profiles.
Project description:Previous data have suggested that B-cell–depletion therapy may induce the settlement of autoreactive long-lived plasma cells (LLPCs) in the spleen of patients with autoimmune cytopenia. To investigate this process, we used the AID-CreERT2-EYFP mouse model to follow PCs engaged in an immune response. Multiplex-PCR at the single-cell level revealed that only a small fraction of splenic PCs had a long-lived signature, whereas PCs present after anti-CD20 antibody treatment appeared more mature, similar to bone-marrow PCs. It suggested that, in addition to a process of selection, a maturation induced upon B-cell depletion drove PCs toward a long-lived program. We showed that BAFF and CD4+ T cells play a major role in PC survival niche, because combining anti-CD20 with anti-BAFF or anti-CD4 antibody greatly reduce the number of splenic PCs. Similar results were obtained in the lupus-prone NZB/W model. These different contributions of soluble and cellular components of the PC niche in the spleen demonstrate that the LLPC expression profile is not cell-intrinsic but largely depends on signals provided by the splenic microenvironment, implying that interfering with these components at the time of B cell depletion might improve the response rate in autoimmune cytopenia.
Project description:The number of peripheral blood (PB) circulating tumor cells (CTCs) predicts risk of transformation in smoldering multiple myeloma (MM) and survival in active MM. Growing evidence suggests that as the tumor progresses and the microenvironment becomes hypoxic, clonal plasma cells (PCs) constantly invade new regions of the bone marrow (BM) through induced systemic recirculation. Of note, the frequency of CTCs is typically low and thus, it could be hypothesized that the dissemination of MM is made by few tumor cells with unique features that induce them to egress the BM and spread the disease through PB. This hypothesis has not been demonstrated because the molecular profile of CTCs in MM has not been investigated. We used gene expression profiling (GEP) arrays to identify gene regulatory networks related to disease dissemination by comparing the molecular profile of CTCs with patient-matched BM clonal PCs.
Project description:The number of peripheral blood (PB) circulating tumor cells (CTCs) predicts risk of transformation in smoldering multiple myeloma (MM) and survival in active MM. Growing evidence suggests that as the tumor progresses and the microenvironment becomes hypoxic, clonal plasma cells (PCs) constantly invade new regions of the bone marrow (BM) through induced systemic recirculation. Of note, the frequency of CTCs is typically low and thus, it could be hypothesized that the dissemination of MM is made by few tumor cells with unique features that induce them to egress the BM and spread the disease through PB. This hypothesis has not been demonstrated because the molecular profile of CTCs in MM has not been investigated. We used gene expression profiling (GEP) arrays to identify gene regulatory networks related to disease dissemination by comparing the molecular profile of CTCs with patient-matched BM clonal PCs.