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Immunity Jun 2022Understanding the drivers and markers of clonally expanding HIV-1-infected CD4 T cells is essential for HIV-1 eradication. We used single-cell ECCITE-seq, which...
Understanding the drivers and markers of clonally expanding HIV-1-infected CD4 T cells is essential for HIV-1 eradication. We used single-cell ECCITE-seq, which captures surface protein expression, cellular transcriptome, HIV-1 RNA, and TCR sequences within the same single cell to track clonal expansion dynamics in longitudinally archived samples from six HIV-1-infected individuals (during viremia and after suppressive antiretroviral therapy) and two uninfected individuals, in unstimulated conditions and after CMV and HIV-1 antigen stimulation. Despite antiretroviral therapy, persistent antigen and TNF responses shaped T cell clonal expansion. HIV-1 resided in Th1-polarized, antigen-responding T cells expressing BCL2 and SERPINB9 that may resist cell death. HIV-1 RNA T cell clones were larger in clone size, established during viremia, persistent after viral suppression, and enriched in GZMB cytotoxic effector memory Th1 cells. Targeting HIV-1-infected cytotoxic CD4 T cells and drivers of clonal expansion provides another direction for HIV-1 eradication.
Topics: CD4-Positive T-Lymphocytes; Clone Cells; HIV Infections; HIV-1; Humans; RNA; Viremia
PubMed: 35320704
DOI: 10.1016/j.immuni.2022.03.004 -
Blood Jul 2015Recent genetic analyses of large populations have revealed that somatic mutations in hematopoietic cells leading to clonal expansion are commonly acquired during human...
Recent genetic analyses of large populations have revealed that somatic mutations in hematopoietic cells leading to clonal expansion are commonly acquired during human aging. Clonally restricted hematopoiesis is associated with an increased risk of subsequent diagnosis of myeloid or lymphoid neoplasia and increased all-cause mortality. Although myelodysplastic syndromes (MDS) are defined by cytopenias, dysplastic morphology of blood and marrow cells, and clonal hematopoiesis, most individuals who acquire clonal hematopoiesis during aging will never develop MDS. Therefore, acquisition of somatic mutations that drive clonal expansion in the absence of cytopenias and dysplastic hematopoiesis can be considered clonal hematopoiesis of indeterminate potential (CHIP), analogous to monoclonal gammopathy of undetermined significance and monoclonal B-cell lymphocytosis, which are precursor states for hematologic neoplasms but are usually benign and do not progress. Because mutations are frequently observed in healthy older persons, detection of an MDS-associated somatic mutation in a cytopenic patient without other evidence of MDS may cause diagnostic uncertainty. Here we discuss the nature and prevalence of CHIP, distinction of this state from MDS, and current areas of uncertainty regarding diagnostic criteria for myeloid malignancies.
Topics: Clonal Evolution; Diagnosis, Differential; Hematologic Neoplasms; Hematopoiesis; Humans; Monoclonal Gammopathy of Undetermined Significance; Mutation; Myelodysplastic Syndromes; Precancerous Conditions
PubMed: 25931582
DOI: 10.1182/blood-2015-03-631747 -
Frontiers in Immunology 2021Paroxysmal Nocturnal Hemoglobinuria (PNH) is a disease as simple as it is complex. PNH patients develop somatic loss-of-function mutations in phosphatidylinositol... (Review)
Review
Paroxysmal Nocturnal Hemoglobinuria (PNH) is a disease as simple as it is complex. PNH patients develop somatic loss-of-function mutations in phosphatidylinositol -acetylglucosaminyltransferase subunit A gene (), required for the biosynthesis of glycosylphosphatidylinositol (GPI) anchors. Ubiquitous in eukaryotes, GPI anchors are a group of conserved glycolipid molecules responsible for attaching nearly 150 distinct proteins to the surface of cell membranes. The loss of two GPI-anchored surface proteins, CD55 and CD59, from red blood cells causes unregulated complement activation and hemolysis in classical PNH disease. In PNH patients, -mutant, GPI (-) hematopoietic cells clonally expand to make up a large portion of patients' blood production, yet mechanisms leading to clonal expansion of GPI (-) cells remain enigmatic. Historical models of PNH in mice and the more recent PNH model in rhesus macaques showed that GPI (-) cells reconstitute near-normal hematopoiesis but have no intrinsic growth advantage and do not clonally expand over time. Landmark studies identified several potential mechanisms which can promote PNH clonal expansion. However, to what extent these contribute to PNH cell selection in patients continues to be a matter of active debate. Recent advancements in disease models and immunologic technologies, together with the growing understanding of autoimmune marrow failure, offer new opportunities to evaluate the mechanisms of clonal expansion in PNH. Here, we critically review published data on PNH cell biology and clonal expansion and highlight limitations and opportunities to further our understanding of the emergence of PNH clones.
Topics: Animals; Autoimmunity; Biomarkers; Clonal Evolution; Cytokines; Disease Management; Disease Models, Animal; Disease Susceptibility; Genetic Predisposition to Disease; Glycosylphosphatidylinositols; Hematopoiesis; Hemoglobinuria, Paroxysmal; Humans; Lymphocyte Subsets; Mutation
PubMed: 35154088
DOI: 10.3389/fimmu.2021.830172 -
Annual Review of Medicine Jan 2023Aging is associated with increased mutational burden in every tissue studied. Occasionally, fitness-increasing mutations will arise, leading to stem cell clonal... (Review)
Review
Aging is associated with increased mutational burden in every tissue studied. Occasionally, fitness-increasing mutations will arise, leading to stem cell clonal expansion. This process occurs in several tissues but has been best studied in blood. Clonal hematopoiesis is associated with an increased risk of blood cancers, such as acute myeloid leukemia, which result if additional cooperating mutations occur. Surprisingly, it is also associated with an increased risk of nonmalignant diseases, such as atherosclerotic cardiovascular disease. This may be due to enhanced inflammation in mutated innate immune cells, which could be targeted clinically with anti-inflammatory drugs. Recent studies have uncovered other factors that predict poor outcomes in patients with clonal hematopoiesis, such as size of the mutant clone, mutated driver genes, and epigenetic aging. Though clonality is inevitable and largely a function of time, recent work has shown that inherited genetic variation can also influence this process. Clonal hematopoiesis provides a paradigm for understanding how age-related changes in tissue stem cell composition and function influence human health.
Topics: Humans; Clonal Hematopoiesis; Hematopoiesis; Hematologic Neoplasms; Aging; Precancerous Conditions; Mutation
PubMed: 36450282
DOI: 10.1146/annurev-med-042921-112347 -
Circulation Jul 2022Immune checkpoint inhibitors (ICIs) are monoclonal antibodies used to activate the immune system against tumor cells. Despite therapeutic benefits, ICIs have the...
BACKGROUND
Immune checkpoint inhibitors (ICIs) are monoclonal antibodies used to activate the immune system against tumor cells. Despite therapeutic benefits, ICIs have the potential to cause immune-related adverse events such as myocarditis, a rare but serious side effect with up to 50% mortality in affected patients. Histologically, patients with ICI myocarditis have lymphocytic infiltrates in the heart, implicating T cell-mediated mechanisms. However, the precise pathological immune subsets and molecular changes in ICI myocarditis are unknown.
METHODS
To identify immune subset(s) associated with ICI myocarditis, we performed time-of-flight mass cytometry on peripheral blood mononuclear cells from 52 individuals: 29 patients with autoimmune adverse events (immune-related adverse events) on ICI, including 8 patients with ICI myocarditis, and 23 healthy control subjects. We also used multiomics single-cell technology to immunophenotype 30 patients/control subjects using single-cell RNA sequencing, single-cell T-cell receptor sequencing, and cellular indexing of transcriptomes and epitopes by sequencing with feature barcoding for surface marker expression confirmation. To correlate between the blood and the heart, we performed single-cell RNA sequencing/T-cell receptor sequencing/cellular indexing of transcriptomes and epitopes by sequencing on MRL/Pdcd1 (Murphy Roths large/programmed death-1-deficient) mice with spontaneous myocarditis.
RESULTS
Using these complementary approaches, we found an expansion of cytotoxic CD8 T effector cells re-expressing CD45RA (Temra CD8 cells) in patients with ICI myocarditis compared with control subjects. T-cell receptor sequencing demonstrated that these CD8 Temra cells were clonally expanded in patients with myocarditis compared with control subjects. Transcriptomic analysis of these Temra CD8 clones confirmed a highly activated and cytotoxic phenotype. Longitudinal study demonstrated progression of these Temra CD8 cells into an exhausted phenotype 2 months after treatment with glucocorticoids. Differential expression analysis demonstrated elevated expression levels of proinflammatory chemokines (CCL5/CCL4/CCL4L2) in the clonally expanded Temra CD8 cells, and ligand receptor analysis demonstrated their interactions with innate immune cells, including monocytes/macrophages, dendritic cells, and neutrophils, as well as the absence of key anti-inflammatory signals. To complement the human study, we performed single-cell RNA sequencing/T-cell receptor sequencing/cellular indexing of transcriptomes and epitopes by sequencing in Pdcd1 mice with spontaneous myocarditis and found analogous expansions of cytotoxic clonal effector CD8 cells in both blood and hearts of such mice compared with controls.
CONCLUSIONS
Clonal cytotoxic Temra CD8 cells are significantly increased in the blood of patients with ICI myocarditis, corresponding to an analogous increase in effector cytotoxic CD8 cells in the blood/hearts of Pdcd1 mice with myocarditis. These expanded effector CD8 cells have unique transcriptional changes, including upregulation of chemokines CCL5/CCL4/CCL4L2, which may serve as attractive diagnostic/therapeutic targets for reducing life-threatening cardiac immune-related adverse events in ICI-treated patients with cancer.
Topics: Animals; Antineoplastic Agents; Antineoplastic Agents, Immunological; Epitopes; Humans; Leukocytes, Mononuclear; Longitudinal Studies; Mice; Myocarditis
PubMed: 35762356
DOI: 10.1161/CIRCULATIONAHA.121.056730 -
The New England Journal of Medicine Dec 2014Cancers arise from multiple acquired mutations, which presumably occur over many years. Early stages in cancer development might be present years before cancers become...
BACKGROUND
Cancers arise from multiple acquired mutations, which presumably occur over many years. Early stages in cancer development might be present years before cancers become clinically apparent.
METHODS
We analyzed data from whole-exome sequencing of DNA in peripheral-blood cells from 12,380 persons, unselected for cancer or hematologic phenotypes. We identified somatic mutations on the basis of unusual allelic fractions. We used data from Swedish national patient registers to follow health outcomes for 2 to 7 years after DNA sampling.
RESULTS
Clonal hematopoiesis with somatic mutations was observed in 10% of persons older than 65 years of age but in only 1% of those younger than 50 years of age. Detectable clonal expansions most frequently involved somatic mutations in three genes (DNMT3A, ASXL1, and TET2) that have previously been implicated in hematologic cancers. Clonal hematopoiesis was a strong risk factor for subsequent hematologic cancer (hazard ratio, 12.9; 95% confidence interval, 5.8 to 28.7). Approximately 42% of hematologic cancers in this cohort arose in persons who had clonality at the time of DNA sampling, more than 6 months before a first diagnosis of cancer. Analysis of bone marrow-biopsy specimens obtained from two patients at the time of diagnosis of acute myeloid leukemia revealed that their cancers arose from the earlier clones.
CONCLUSIONS
Clonal hematopoiesis with somatic mutations is readily detected by means of DNA sequencing, is increasingly common as people age, and is associated with increased risks of hematologic cancer and death. A subset of the genes that are mutated in patients with myeloid cancers is frequently mutated in apparently healthy persons; these mutations may represent characteristic early events in the development of hematologic cancers. (Funded by the National Human Genome Research Institute and others.).
Topics: Adult; Age Factors; Aged; Aged, 80 and over; Blood; Cell Transformation, Neoplastic; Clone Cells; DNA Mutational Analysis; Exome; Hematologic Neoplasms; Hematopoiesis; Hematopoietic Stem Cells; Humans; Middle Aged; Mutation; Risk Factors; Young Adult
PubMed: 25426838
DOI: 10.1056/NEJMoa1409405 -
Nature Apr 2011Genomic analysis provides insights into the role of copy number variation in disease, but most methods are not designed to resolve mixed populations of cells. In...
Genomic analysis provides insights into the role of copy number variation in disease, but most methods are not designed to resolve mixed populations of cells. In tumours, where genetic heterogeneity is common, very important information may be lost that would be useful for reconstructing evolutionary history. Here we show that with flow-sorted nuclei, whole genome amplification and next generation sequencing we can accurately quantify genomic copy number within an individual nucleus. We apply single-nucleus sequencing to investigate tumour population structure and evolution in two human breast cancer cases. Analysis of 100 single cells from a polygenomic tumour revealed three distinct clonal subpopulations that probably represent sequential clonal expansions. Additional analysis of 100 single cells from a monogenomic primary tumour and its liver metastasis indicated that a single clonal expansion formed the primary tumour and seeded the metastasis. In both primary tumours, we also identified an unexpectedly abundant subpopulation of genetically diverse 'pseudodiploid' cells that do not travel to the metastatic site. In contrast to gradual models of tumour progression, our data indicate that tumours grow by punctuated clonal expansions with few persistent intermediates.
Topics: Breast Neoplasms; Carcinoma, Ductal, Breast; Chromosome Breakpoints; Clone Cells; Diploidy; Disease Progression; Evolution, Molecular; Female; Flow Cytometry; Genetic Heterogeneity; Genome, Human; Genomics; Humans; Liver Neoplasms; Loss of Heterozygosity; Sequence Analysis, DNA; Single-Cell Analysis
PubMed: 21399628
DOI: 10.1038/nature09807 -
Cell Jan 2020Repeated exposure to pathogens or their antigens triggers anamnestic antibody responses that are higher in magnitude and affinity than the primary response. These...
Repeated exposure to pathogens or their antigens triggers anamnestic antibody responses that are higher in magnitude and affinity than the primary response. These involve reengagement of memory B cell (MBC) clones, the diversity and specificity of which determine the breadth and effectiveness of the ensuing antibody response. Using prime-boost models in mice, we find that secondary responses are characterized by a clonality bottleneck that restricts the engagement of the large diversity of MBC clones generated by priming. Rediversification of mutated MBCs is infrequent within secondary germinal centers (GCs), which instead consist predominantly of B cells without prior GC experience or detectable clonal expansion. Few MBC clones, generally derived from higher-affinity germline precursors, account for the majority of secondary antibody responses, while most primary-derived clonal diversity is not reengaged detectably by boosting. Understanding how to counter this bottleneck may improve our ability to elicit antibodies to non-immunodominant epitopes by vaccination.
Topics: Adaptive Immunity; Animals; Antibody Formation; Antigens; B-Lymphocytes; CHO Cells; Cell Line; Cricetulus; Female; Germinal Center; Humans; Immunologic Memory; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Models, Animal
PubMed: 31866068
DOI: 10.1016/j.cell.2019.11.032 -
Experimental Hematology Mar 2020Clonal hematopoiesis is a common premalignant condition defined by the abnormal expansion of clonally derived hematopoietic stem cells carrying somatic mutations in... (Review)
Review
Clonal hematopoiesis is a common premalignant condition defined by the abnormal expansion of clonally derived hematopoietic stem cells carrying somatic mutations in leukemia-associated genes. Apart from increasing age, this phenomenon occurs with higher frequency in individuals with lymphoid or solid tumors and is associated with exposures to genotoxic stress. Clonal hematopoiesis in this context confers a greater risk for developing therapy-related myeloid neoplasms and appears to contribute to adverse cancer-related survival through a variety of potential mechanisms. These include alterations of the bone marrow microenvironment, inflammatory changes in clonal effector cells and modulation of immune responses. Understanding how clonal hematopoiesis drives therapy-related myeloid neoplasm initiation and interactions with non-myeloid malignancies will inform screening and surveillance approaches and suggest targeted therapies in this vulnerable population. Here, we examine the clinical implications of clonal hematopoiesis in the cancer setting and discuss potential strategies to mitigate the adverse consequences of clonal expansion.
Topics: Clonal Evolution; DNA Damage; Hematologic Neoplasms; Hematopoiesis; Humans; Myeloproliferative Disorders
PubMed: 32044376
DOI: 10.1016/j.exphem.2020.02.001 -
Cell May 2012Cancer evolves dynamically as clonal expansions supersede one another driven by shifting selective pressures, mutational processes, and disrupted cancer genes. These...
Cancer evolves dynamically as clonal expansions supersede one another driven by shifting selective pressures, mutational processes, and disrupted cancer genes. These processes mark the genome, such that a cancer's life history is encrypted in the somatic mutations present. We developed algorithms to decipher this narrative and applied them to 21 breast cancers. Mutational processes evolve across a cancer's lifespan, with many emerging late but contributing extensive genetic variation. Subclonal diversification is prominent, and most mutations are found in just a fraction of tumor cells. Every tumor has a dominant subclonal lineage, representing more than 50% of tumor cells. Minimal expansion of these subclones occurs until many hundreds to thousands of mutations have accumulated, implying the existence of long-lived, quiescent cell lineages capable of substantial proliferation upon acquisition of enabling genomic changes. Expansion of the dominant subclone to an appreciable mass may therefore represent the final rate-limiting step in a breast cancer's development, triggering diagnosis.
Topics: Algorithms; Breast Neoplasms; Cell Transformation, Neoplastic; Chromosome Aberrations; Clonal Evolution; Female; Humans; Mutation; Point Mutation
PubMed: 22608083
DOI: 10.1016/j.cell.2012.04.023