SEARCH · PubMed Health

Results for “clonality”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 127 records · Page 7Linked to original sources

PubMed1999

Enhanced sensitivity with a novel TCRgamma PCR assay for clonality studies in 569 formalin-fixed, paraffin-embedded (FFPE) cases.

BACKGROUND: Clonal rearrangement of genes encoding the immunoglobulins (Ig) and T-cell antigen receptors (TCR) are considered to be useful markers for the diagnosis of lymphoma and for determining the clonal origins of B- and T-cell populations in lymphoid neoplasms. METHODS AND RESULTS: Polymerase chain reaction-based clonality assays for TCRgamma, TCRbeta, and immunoglobulin (Ig) heavy chain (IgH) gene rearrangements were evaluated for diagnostic sensitivity and specificity in 569 formalin-fixed, paraffin-embedded (FFPE) tissues. Combined TCRbeta and TCRgamma assays enhanced the routine detection of TCR clonality to 90% of all peripheral T-cell lymphoma (PTCL) cases. IgH clonality was detected in 59% of 241 peripheral B-cell lymphoma (BCL) cases and 6% of 169 PTCL cases. Of 452 lymphomas, 5% could not be classified phenotypically as B or T lineage after immunohistochemical and clonality studies. Of all BCL cases analyzed, 24% had detectable TCRbeta and/or TCRgamma clonality. Of these BCL with biclonal results, 47% were extranodal lymphomas from skin and various tissues. CONCLUSIONS: Clonality assays were useful for distinguishing reactive or benign lymph nodes from neoplastic lymphoid infiltrates in most cases. The inclusion of TCRb and TCRg assays in the assessment of lymphomas results in a significant increase in the sensitivity of clonality detection, but is of limited utility in assessing the T- or B-cell phenotype of the tumor.

B-Lymphocytes↗

PubMed2000

Lack of co-ordinate expression of the alpha1(I) and alpha1(III) procollagen genes in fibroblast clonal cultures.

BACKGROUND: Several extracellular matrix genes, most notably alpha1(I) and alpha1(III) procollagen, are reported to be co-ordinately expressed in cultures of dermal fibroblasts. However, it remains unclear whether the expression of these genes is truly co-ordinate or whether it may be the result of averaging the phenotypic expression of different fibroblast subpopulations present within each culture. Objectives To determine by Northern analysis the correlation between alpha1(I) and alpha1(III) procollagen mRNA levels in clonal populations of human dermal fibroblasts. METHODS: As previously described, clonal cultures were derived from parent strains of human dermal fibroblasts by a microscopically controlled dilution technique and by stimulation of single cells with low oxygen tension in the early phases of clonal growth. RESULTS: In agreement with previous reports, we found that baseline steady-state levels of alpha1(I) procollagen mRNA were co-ordinately regulated with the alpha1(III) procollagen mRNA in 26 parent strains (r = 0. 9003; P < 0.0001). However, this close correlation between the expression of these two procollagen chains was absent in a total of 40 unselected clonal strains derived from four of the parent cultures (r = 0.5745; P < 0.0001). Moreover, this intrachain heterogeneity in alpha1(I) and alpha1(III) procollagen mRNA levels in clonal cultures was statistically significant from that measured in parent strains (P = 0.0016). CONCLUSIONS: alpha1(I) and alpha1(III) procollagen mRNA levels in clonal cultures do not show the tight co-ordinate regulation observed in non-clonal cultures, suggesting that these two genes operate under different sets of regulatory controls. This clonal heterogeneity may provide additional flexibility to the process of tissue repair and fibroblast clonal expansion.

Adult↗

PubMed2001

Interleukin 4 content in chronic lymphocytic leukaemia (CLL) B cells and blood CD8+ T cells from B-CLL patients: impact on clonal B-cell apoptosis.

B-chronic lymphocytic leukaemia (CLL) clonal B cells are characterized by resistance to apoptosis. We evaluated clonal B cells and blood T cells for interleukin 4 (IL-4) content as IL-4 is able to increase CLL cell resistance to apoptosis. The content of IL-4 in CD8+ T cells of CLL patients (n = 9) ranged from 37% to 63% of the total CD8+ T cells (mean level of 49% +/- 3.4) compared with a range of 5-10% for control CD8+ T cells. Clonal B cells positive for cytoplasmic IL-4 ranged from 1% to 97% (mean value 57.8 +/- 6.9%). CD8+ T cells and clonal B cells secreted detectable levels of IL-4, but only clonal CLL B cells (n = 4) secreted IL-4 in association with increasing cell numbers. Fludarabine (F-ara-AMP, 0.1-100 micromol/ml) was able to downregulate the IL-4 content of CD8+ T cells, but not clonal B-cell IL-4. Culture supernatant from CLL CD8+ T cells decreased the spontaneous apoptotic rate of clonal B cells that was reversed with anti-IL-4 and soluble IL-4 receptor. These findings show that IL-4 is present in the microenvironment of B-CLL. In addition, use of agents that can interfere with IL-4 presentation to clonal B cells can be effective in increasing clonal B-cell apoptosis.

Antineoplastic Agents↗

PubMed1990

Genetic variation in clonal vertebrates detected by simple-sequence DNA fingerprinting.

The measurement of clonal heterogeneity is central to understanding the evolutionary and population genetics of the roughly 50 species of vertebrates that lack effective genetic recombination. Simple-sequence DNA fingerprinting with oligonucleotide probes (CAC)5 and (GACA)4 is a sensitive and efficient means of detecting this heterogeneity in natural populations of two clonal fishes, Poecilia formosa, an apomictic unisexual, and Rivulus marmoratus, a selfing hermaphrodite. The fingerprints are clonally stable for at least three generations. The technique clearly differentiates allozymically identical laboratory lines of R. marmoratus that were previously distinguishable only by histocompatibility analysis. The technique also reveals apparent cases of shifts in clonal composition of a natural population of each species. Clonal variation in most natural populations is quite high. For example, a sample of 19 specimens of P. formosa from one station on the Rio Soto la Marina contained 16 clones (average clonal frequency = 0.07). This level of clonal diversity implies that mutation, subsequent to the founding of clonal lineages, is an important source of variation in these populations. It also suggests that chance (sampling error) has a previously unappreciated role in determining the clonal composition of populations even though some of the clones may be divergent in biologically significant features.

Animals↗

PubMed1994

Detection of clonal T-cell receptor gamma gene rearrangements in early mycosis fungoides/Sezary syndrome by polymerase chain reaction and denaturing gradient gel electrophoresis (PCR/DGGE).

We used a gene amplification strategy to analyze T-cell receptor (TCR) gene rearrangements in 185 specimens, including mycosis fungoides/Sezary syndrome (MF/SS), other cutaneous neoplasms, inflammatory dermatoses, reactive lymphoid tissues, and normal skin. Genomic DNA was extracted from lesional tissues and rearrangements of the TCR-gamma chain gene were amplified using the polymerase chain reaction (PCR) with primers specific for rearrangements involving V gamma 1-8 or V gamma 9 gene segments. The resulting PCR products were then separated according to their nucleotide sequence as well as size by denaturing gradient gel electrophoresis (DGGE). Dominant clonal TCR-gamma gene rearrangements were detected in 61 of 68 MF/SS cases by PCR/DGGE. This sensitivity of 90% compared to a sensitivity of only 59% when dominant clonality was sought in 17 of these same cases by Southern blot analysis of TCR-beta gene rearrangements. This difference in sensitivity was greatest in early, minimally infiltrated skin lesions. PCR/DGGE was also more sensitive than Southern blot analysis for detecting peripheral blood involvement in two cases of early MF. Among 12 additional specimens of suspected MF/SS, nine (75%) showed clonal TCR-gamma gene rearrangements by PCR/DGGE including six of eight cases with a previously confirmed diagnosis of MF/SS and three of four cases without prior known MF/SS. Among 105 non-MF/SS specimens, dominant TCR-gamma gene rearrangements were detected in only six cases (6%). Four were diagnosed as chronic dermatitis and two were diagnosed as cutaneous lymphoid hyperplasia. We conclude that the large majority of MF/SS cases, including patch phase disease, possess dominant clonal TCR-gamma gene rearrangements. PCR/DGGE is more sensitive than Southern blot analysis for detecting dominant clonality and staging disease in patients with a confirmed diagnosis of MF/SS. However, because PCR/DGGE is sensitive enough to detect dominant TCR-gamma gene rearrangements in a subset of patients with chronic dermatitis, it cannot be used as the sole criterion for establishing a diagnosis of T-cell lymphoma. As with other molecular biologic clonality assays, clinicopathologic correlation is essential. Nevertheless, the detection of dominant clonality in some cases of histologically nonspecific dermatitis allows the identification of a previously unrecognized subset of patients, i.e., those with "clonal dermatitis." It will be important to determine the long-term risk of MF/SS among these patients because our study indicated that MF/SS can sometimes present with lesions indistinguishable from clonal dermatitis.

Base Sequence↗

PubMed1993

Detection of clonal platelet antibodies in immunologically-mediated thrombocytopenias: association with circulating clonal/oligoclonal B cells.

Aware that T and B cells in autoimmune thrombocytopenia are abnormal, including the existence of clonal B cell populations, we sought to characterize this clonal phenomenon in various immunological thrombocytopenias using platelet antibody light chain analysis, flow cytometry, Southern blot analysis, and PCR. Using a monoclonal antibody-antigen capture ELISA, we analysed sera from 21 of 26 patients with autoimmune, alloimmune, or drug-induced immunological thrombocytopenia for the light chain phenotypes of their platelet antibodies. Alloantibodies and drug-dependent antibodies from four and 14 patients, respectively, were found that expressed a predominant type of light chain, suggesting that these platelet-reactive antibodies were monoclonal or oligoclonal in nature. 14 of the 26 patients were available for light chain B cell phenotyping studies. Of these 14 patients, thrombocytopenia was due to autoimmunity in two, drug-induced immunity in four, and alloimmunity in eight. We detected clonal populations of B cells in all 14 patients by flow cytometry. Although six of these latter patients possessed platelet antibodies with clonal characteristics, light chain phenotypes of antibodies in five patients were opposite to those of their B cells. Eight of these patients were further examined for immunoglobulin gene rearrangement using Southern and/or polymerase chain reaction analysis. In all eight patients we detected clonal or oligoclonal B cell populations. Only two of these patients had malignancies (chronic lymphocytic leukaemia) that would be expected to have detectable clonal B cells, and thus the mechanism for clonal expansion in the other six patients did not appear to be related to an obvious neoplastic process. Prior to these studies, detection of clonal B cells in thrombocytopenic patients without known malignancies was limited to individuals with autoimmune thrombocytopenia, prompting the speculation that this particular autoimmune disorder arises from B cell dysregulation, rather than from expansion of specific autoantibody producing B cell clones. In contrast, the current studies provide evidence that clonal B cells are common to patients with any form of immunologically-mediated thrombocytopenia. Moreover, the majority of the platelet antibodies (86%) present in these disorders exhibited monoclonal characteristics in that there was an apparent restriction in light chain usage.

Adult↗

PubMed1993

Clonal remissions in acute myeloid leukaemia are commonly associated with features of trilineage myelodysplasia during remission.

Clonal haemopoiesis has previously been demonstrated in some 30% of patients in remission of acute myeloid leukaemia (AML). Whilst a 'clonal remission' in many such patients may represent a skewed X-chromosome inactivation pattern in haemopoietic cells, its relationship to an underlying preleukaemic state remains uncertain. We therefore analysed the clonal status of 48 female patients in remission of AML using X-chromosome linked restriction fragment length polymorphisms (RFLPs) within the X-linked PGK and HPRT genes and the DXS255 (M27 beta) locus, and carried out in conjunction a detailed study of the morphological and karyotypic features of the patients' bone marrows. During remission, 35 patients (73%) with AML demonstrated nonclonal haemopoiesis, and their bone marrows were morphologically normal. Remission haemopoietic tissue in nine cases (19%) showed a skewed X-chromosome inactivation pattern and remission bone marrows in these patients had features of trilineage myelodysplasia (TMDS), with seven having similar features at presentation. Analysis of constitutional DNA showed a non-clonal pattern in seven of these patients, but was unsuccessful in two cases. These nine patients with post-chemotherapy TMDS were considered to have true clonal haemopoiesis. Four patients (8%) with a skewed X-chromosome inactivation pattern had normal remission bone marrows. Analysis of constitutional DNA showed a skewed pattern in two of these patients, but was unsuccessful in two cases. Cytogenetic investigation during remission in the nine patients with TMDS showed a normal karyotype in four cases and the acquisition of new karyotypic abnormalities in three cases. In contrast, 10 female patients in remission of de novo acute lymphoblastic leukaemia (ALL) were shown to have non-clonal haemopoiesis. We conclude that the majority of patients with AML who achieve remission after cytoreductive chemotherapy have non-clonal haemopoiesis, and when clonal remissions are observed these are commonly associated with the development of trilineage myelodysplasia in the bone marrow, with or without karyotypic abnormalities. True clonal remission in association with morphologically normal haemopoiesis is a rare entity, the significance and frequency of which remain uncertain.

Aged↗

PubMed2007

Correlated evolution of self-incompatibility and clonal reproduction in Solanum (Solanaceae).

It has been suggested that clonality provides reproductive assurance in cross-fertilizing species subject to pollen limitation, relieving one of the main selective pressures favoring the evolution of self-fertilization. According to this hypothesis, cross-fertilizing species subject to pollen limitation should often be clonal. Here, we investigated the association between clonality and a genetic mechanism enforcing outcrossing, self-incompatibility, in Solanum (Solanaceae). We collected self-incompatibility and clonality information on 87 species, and looked for an association between these two traits. To account for the contribution of shared evolutionary history to this association, we incorporated phylogenetic information from chloroplast (NADH dehydrogenase subunit F) sequence data. We found that self-incompatibility is strongly associated with clonal reproduction: all self-incompatible species reproduce clonally, while the absence of clonality is widespread among self-compatible taxa. The observed correlation persists after taking into account shared phylogenetic history, assumptions about the evolutionary history of self-incompatibility, uncertainty associated with phylogeny estimation, and associations with life history (annual/perennial). Our results are consistent with the hypothesis that clonality provides reproductive assurance, and suggest that the consequences of clonal growth in the evolution of plant reproductive strategies may be more significant than previously thought.

Biological Evolution↗

PubMed2004

Clonal B cell populations in a minority of patients with Hashimoto's thyroiditis.

BACKGROUND: Hashimoto's thyroiditis (HT) is a risk factor for thyroid lymphoma, and clonal B cell populations in HT support this link. The literature on B cell clonality in HT is controversial. AIMS: To identify clonal B cell populations in HT and to assess their usefulness in differentiating HT from mucosa associated lymphoid tissue (MALT) lymphoma and predicting future development of lymphoma. METHODS: DNA from formalin fixed, paraffin wax embedded blocks of thyroid specimens from 10 patients with HT and two thyroid MALT lymphomas was analysed for B cell clonality by seminested polymerase chain reaction (PCR) using FRIII/LJH and FRIII/VLJH primers to amplify the IgH gene VDJ region. In one case, PCR products were sequenced. Immunohistochemistry was performed by labelled streptavidin-biotin technique using antibodies to: CD45, CD45RO, CD3, CD20, and cytokeratin. RESULTS: The histopathological and clinical findings were characteristic of HT. Clonal bands were seen in three and a polyclonal smear pattern was seen in seven cases. The clonal bands in HT were associated with a background smear, and could not be reproduced from other blocks from the same case or from deeper sections of the same block. The clonal bands in thyroid lymphomas were not associated with a background smear and were reproducible. None of the patients with clonal B cells has developed malignant lymphoma during a follow up of 10-13 years. CONCLUSIONS: B cell clonal bands in HT have different features from those in lymphoma (non-pure and non-reproducible) and do not predict future development of lymphoma.

Adult↗

PubMed2000

Lymphokine-activated killer (LAK) cells and cytokines synergize to kill clonal cells in acute myeloid leukemia (AML) in vitro.

We studied the influence of autologous lymphokine-activated-killer (LAK) cells on the survival of clonal and CD34-positive bone marrow (BM) cells from patients with acute myeloid leukemia (AML) in a coculture assay in vitro. (1) LAK cells were grown in the presence of IL-2, in some cases additionally with IL-6. (2) These cytotoxic cells were cocultured with (untreated or cytokine pretreated) AML-BM cells obtained at different stages of the disease. Therefore BM cells were (a) either frozen in liquid nitrogen or (b) precultured for 14 days with cytokines: IL-1beta, IL-3, IL-6, erythropoietin (EPO), stem cell factor (SCF) with ('Cytok1') or without granulocyte macrophage colony stimulating factor (GM-CSF) ('Cytok2') or with no added cytokines ('ISC/FCS') as a control. (3) Southern blot analysis was used to detect clonal BM cells. At diagnosis, 76 of 151 cases (50%) studied showed clonal gene rearrangements in marker genes. (4) Southern blot analysis and flow cytometry were used to compare the amount of clonal and CD34 positive BM cells before and after coculture procedures. Coculture experiments with untreated BM and autologous LAK cells led to a reduction of clonal cells in 2 of 5 cases at diagnosis, in 11 of 17 BM samples in complete remission but not in the one case studied at relapse. Similar results were found if precultured AML cells (with or without cytokines) were cocultivated with LAK cells. However the cytotoxic effect of LAK cells was more pronounced if cytokines (especially GM-CSF and SCF) were comprised. Our data indicate, that (1) clonality in AML can be demonstrated by Southern blot analysis; (2) CD34 positive cells in AML are clonal, gene rearranged cells; (3) clonal cell populations persist in BM during complete remission and relapse in most of the patients; (4) incubation of AML-BM cells with LAK cells lead to a reduction of clonal, rearranged cells in 11 of 17 AML cases in complete remission, but only in 2 of 6 cases at diagnosis or relapse; (5) AML cells can be sensitized to theLAK cell treatment by preincubation of AML-BM cells with cytokines (IL-1beta, IL-3, IL-6, SCF, EPO and GM-CSF) or by adding SCF to the coculture conditions. Southern blot analysis and flow cytometry are appropriate methods to detect and quantify leukemic disease. Cytokines and LAK cells synergize to kill AML blasts in vitro. This is a feasible approach to immunotherapy of AML and merits further investigations.

Acute Disease↗

PubMed2002

Discrimination of polycythemias and thrombocytoses by novel, simple, accurate clonality assays and comparison with PRV-1 expression and BFU-E response to erythropoietin.

Essential thrombocythemia (ET) and polycythemia vera (PV) are clonal myeloproliferative disorders that are often difficult to distinguish from other causes of elevated blood cell counts. Assays that could reliably detect clonal hematopoiesis would therefore be extremely valuable for diagnosis. We previously reported 3 X-chromosome transcription-based clonality assays (TCAs) involving the G6PD, IDS, and MPP1 genes, which together were informative in about 65% of female subjects. To increase our ability to detect clonality, we developed simple TCA for detecting the transcripts of 2 additional X-chromosome genes: Bruton tyrosine kinase (BTK) and 4-and-a-half LIM domain 1 (FHL1). The combination of TCA established the presence or absence of clonal hematopoiesis in about 90% of female subjects. We show that both genes are subject to X-chromosome inactivation and are polymorphic in all major US ethnic groups. The 5 TCAs were used to examine clonality in 46 female patients along with assays for erythropoietin-independent erythroid colonies (EECs) and granulocyte PRV-1 mRNA levels to discriminate polycythemias and thrombocytoses. Of these, all 19 patients with familial polycythemia or thrombocytosis had polyclonal hematopoiesis, whereas 22 of 26 patients with clinical evidence of myeloproliferative disorder and 1 patient with clinically obscure polycythemia were clonal. Interestingly, interferon alpha therapy in 2 patients with PV was associated with reversion of clonal to polyclonal hematopoiesis. EECs were observed in 14 of 14 patients with PV and 4 of 12 with ET, and increased granulocyte PRV-1 mRNA levels were found in 9 of 13 patients with PV and 2 of 12 with ET. Thus, these novel clonality assays are useful in the diagnosis and follow-up of polycythemic conditions and disorders with increased platelet levels.

Adolescent↗

PubMed1987

Clonal lifespan of Paramecium tetraurelia: effect of selection on its extension and use of fissions for its determination.

Paramecia cells, like human diploid cells cultured in vitro, provide a useful model system for understanding the mechanism that limits division potential. The reported maxima of the clonal lifespan of Paramecium tetraurelia fall into two ranges: from 220 to 258 fissions and from 310 to 325 fissions. We found that neither the selection of vigorous lines nor the cryptic occurrence of autogamy offers a plausible explanation for the much longer lifespans in the latter range. We found the sporadic occurrence of very long clonal lifespans, such as 330 fissions, without selection and autogamy. Selection, which was evaluated by using different methods to maintain lines, had little effect on the extension of the maximal clonal lifespan, whereas it did have a marked effect on the extension of the mean clonal lifespan. Autogamy, which was checked with two closely linked marker genes, was frequent, but only during the period when lines were terminating. Statistical analysis on the mean clonal lifespans of two groups of subclones cultured at 25 and 20 degrees C or with rich and poor nutrition showed that the clonal lifespan that was fission-related under favourable conditions tended to dissociate from fissions under less favourable conditions. We discuss mechanisms that determine the clonal lifespan, programmed events contributing to the maximum clonal lifespan and random events contributing to the mean clonal lifespan.

Animals↗

PubMed1987

Effect of recombinant monokines, lymphokines, and other agents on clonal proliferation of human lung cancer cell lines.

The modulation of clonal growth of cells of 15 human lung cancer lines was examined by coculture with different recombinant lymphokines, monokines, and several agents which induce differentiation in other malignant cell systems. Recombinant human tumor necrosis factor alpha (TNF) was inhibitory to all non-small cell lung cancer cell lines with a 50% effective dose of clonal inhibition (ED50) in the range of 30-2000 units/ml. Two representative squamous lines (SK-MES and P3) had 150 to 250 high affinity (Kd approximately equal to pM) cell surface TNF receptors. In contrast, clonal growth of small cell lung cancer lines was not inhibited by TNF, and two representative lines (H69c and R592) expressed negligible cell surface TNF receptors. Recombinant alpha, beta, and gamma interferons (4000 units/ml) each inhibited greater than or equal to 30% clonal growth of more than 50% of the non-small cell lung cancer lines. TNF (100-1000 units/ml) in combination with gamma-interferon was synergistic in the inhibition of clonal growth of these cells. Further studies showed that synergism of clonal inhibition occurred even when the cells were initially exposed to gamma-interferon, washed, and plated in soft agar with TNF. All-trans-retinoic acid (ED50, 5 X 10(-7)-10(-6) M), dimethyl sulfoxide (ED50, 1.2-1.6%), and 12-O-tetradecanoylphorbol-13-acetate (ED50, 5 X 10(-8)-10(-10) M) inhibited clonal proliferation of 7 of 9, 7 of 9, and 8 of 9 non-small cell lung cancer lines, respectively. In contrast, clonal proliferation of cells of small cell lung cancer lines was decreased only slightly at almost all concentrations of each of the agents. Interleukin-1 and -2 and granulocyte-monocyte colony-stimulating factor had no effect on the clonal growth of any of the lung cancer lines. Our results suggest that TNF in combination with gamma-interferon may be therapeutically active for some patients with non-small cell lung cancer, but small cell lung cancer probably will be unresponsive to all the agents that we examined.

Calcitriol↗

PubMed1995

Analysis of meningiomas by methylation- and transcription-based clonality assays.

The clonal derivation of tumors can be determined by X chromosome inactivation analysis based on differential expression of genes or differential methylation of cytosine residues in CpG islands near polymorphic loci. In this report, we compared a transcription-based RNA analysis with a methylation-based DNA assay to determine clonality of meningiomas. Both clonality assays use PCR-based analysis at the hunan androgen-receptor gene (HUMARA) on the X chromosome. Among 23 meningiomas from female patients, 19 were informative heterozygotes at this locus (83%). The patterns of X chromosome inactivation in four patients were extremely skewed towards one allele in blood (unequal Lyonization), which precluded clonality determination in the tumor samples. Concordant clonality results with methylation- and transcription-based clonality assays were demonstrated in 9 of 13 informative tumors expressing the androgen receptor. Seven meningiomas were monoclonal, but surprisingly, two pathologically documented cases of meningiomas were polyclonal. There was disparity in 4 of 13 tumor specimens that were polyclonal by the methylation-based assay but monoclonal by the transcription assay. Clonality examination of these tumors by the methylation-based phosphoglycerate kinase assay provided identical results to the methylation-based analysis at the HUMARA locus. In addition, loss of heterozygosity (LOH) studies of chromosome 22, which is frequently deleted in meningiomas, showed that four of four informative samples of the six polyclonal tumors had partial LOH in tumor tissues. However, complete LOH was observed in primary cultured cells, which were also monoclonal by the methylation assay. Taken together, these data suggest that the disparity of the two assays in these four cases may be due to differences in the level of expression of the androgen receptor gene in tumors. Therefore, we conclude that: (a) clonal derivation of meningiomas determined by both transcription- and methylation-based clonality assays are in full agreement in many (9 of 13) but not all cases (4 of 13); and (b) most meningiomas (9 of 15) are monoclonal in origin, whereas some meningioma samples (6 of 15) are polyclonal or may contain heterogeneous components.

Adult↗

PubMed1998

Predictive value of clonality assays in patients with non-Hodgkin's lymphoma undergoing autologous bone marrow transplant: a single institution study.

Recent studies have documented an increased risk of therapy-related myelodysplastic syndrome or acute myelogenous leukemia (t-MDS/AML) after autologous bone marrow transplant (ABMT) for non-Hodgkin's lymphoma (NHL). To develop methods to identify patients at risk for this complication, we have investigated the predictive value of clonal bone marrow (BM) hematopoiesis for the development of t-MDS/AML, as defined by an X-inactivation based clonality assay at the human androgen receptor locus (HUMARA), in a group of patients undergoing ABMT for NHL from a single institution (Dana-Farber Cancer Institute, Boston, MA). One hundred four female patients were analyzed. At the time of ABMT, the prevalence of polyclonal hematopoiesis was 77% (80/104), of skewed X-inactivation pattern (XIP) was 20% (21/104), and of clonal hematopoiesis was 3% (3/104). To determine the predictive value of clonality for the development of t-MDS/AML, a subgroup of 78 patients with at least 18 months follow-up was analyzed. As defined by the HUMARA assay, 53 of 78 patients had persistent polyclonal hematopoiesis, 15 of 78 had skewed XIP, and 10 of 78 (13.5%) either had clonal hematopoiesis at the time of ABMT or developed clonal hematopoiesis after ABMT. t-MDS/AML developed in 2 of 53 patients with polyclonal hematopoiesis and in 4 of 10 with clonal hematopoiesis. We conclude that a significant proportion of patients have clonal hematopoiesis at the time of ABMT and that clonal hematopoiesis, as detected by the HUMARA assay, is predictive of the development of t-MDS/AML (P = .004).

Adult↗

PubMed1998

Clonal analysis of hematopoietic cells using a novel polymorphic site of the X chromosome.

Clonality of hematopoietic cells on a smale scale (nanogram amounts of DNA) can be detected by X-chromosome inactivation using the polymerase chain reaction (PCR). The human androgen-receptor gene (HUMARA) has a polymorphic short tandem repeat (STR), and has generally been used for clonality analysis since heterozygosity for the gene occurs in 90% of caucasian females. We examined heterozygosity of the STR on HUMARA in 110 Japanese females and found heterozygosity in 74 of 110 (67%). To examine for hematologic clonality in females with HUMARA homozygosity, we used a primer specific for a novel polymorphic STR site between DXS15 and DXS134 (DXS15-134) on Xq28. Heterozygosity for this site was found in 50 of 110 females (46%). Clonality of the hematopoietic cells was detected in 91 of 110 females (83%) using PCR of either the STR sites on HUMARA or DXS15-134. The X-inactivation patterns using PCR of DXS15-134 corresponded exactly with those obtained using PCR of HUMARA in 18 females who were heterozygous for both DXS15-134 and HUMARA. Using PCR of DXS15-134, we examined the clonality of bone marrow cells separated by flow cytometry in a patient with erythroleukemia (M6). Clonality was found not only in myeloid lineage cells but also in B lymphocytes. The clonality assay for DXS15-134 may be useful to assess for clonality of hematopoietic cells in the Japanese population, when combined with the HUMARA assay.

Base Sequence↗

PubMed2000

Cytogenetic and Southern blot analysis to demonstrate clonality and to estimate prognosis in patients with myelodysplastic syndromes.

We examined the bone marrow of 109 patients with myelodysplastic syndrome (MDS) at the time of diagnosis and during the course of the disease by means of Southern blot analysis and/or cytogenetic studies to detect and evaluate clonal markers, their implications for the prognosis of the disease, and the response to treatment. The patients either were enrolled in an EORTC study and received low-dose Ara-C with (n=31) or without (n=21) growth factors, according to the study protocol, or were treated supportively (only one patient received regular chemotherapy for concomitant lymphoma). Full or at least partial remission was achieved by 34% of the treated patients (n=54). In 57% (53 of 93) of all patients a clonal marker of either kind was detected by Southern blot analysis and/or cytogenetic examination. Clonal chromosomal aberrations were found in 45% (35 of 77) of the cases examined at diagnosis, with solitary del(5q) aberrations occurring in 10% of the cases and complex aberrations in 18%, trisomy 8 or monosomy 7 being a frequent finding. Of all patients, 49% (28 of 57) were characterized by one or more gene rearrangements (e.g., Ig-JH, TcR-beta, M-bcr, GM-CSF, G-CSF, or IL-3) at diagnosis. In five of 21 cases (24%) studied in hematological remission of the disease chromosomal aberrations were still detectable, and in seven of 23 (30%) a gene rearrangement persisted. We also found six cases with multiple clones exhibiting different susceptibilities to treatment and thereby indicating the oligoclonal character of this disease. Clinical evaluation revealed that the prognosis of the respective patients was directly related to the particular clonal markers detected at diagnosis: Risk groups were subdivided according to the karyotypes, with a solitary del(5q) aberration meaning a favorable, a normal karyotype an intermediate, solitary aberrations without del(5q) a poor, and complex karyotypes a very poor prognosis. We showed that densitometry helps to increase the sensitivity of Southern blot analysis by quantifying the amount of altered DNA, which often increases shortly before or at progression of MDS. Overall, there was a high level of concordance of both clonality examinations with the clinical course of the disease and the response rate. Therefore, we recommend cytogenetic studies and Southern blot analysis to detect clonal markers at diagnosis of MDS, to detect oligoclonality and clonal evolution, or to quantify the amount of clonal DNA, which appears to be a sensitive tool for evaluating the prognosis and response to therapy in MDS.

Aged↗

PubMed1994

Plasma interleukin-6 and C-reactive protein levels in reactive versus clonal thrombocytosis.

PURPOSE: Evaluate the discriminatory value of plasma interleukin-6 or C-reactive protein levels in clonal thrombocytosis compared with those in reactive thrombocytosis. PATIENTS AND METHODS: A comparative analysis of quantitatively measured laboratory values in a prospectively studied group of consecutive patients. The setting was a tertiary referral center consisting of two hospitals and an outpatient clinic. Plasma interleukin-6 and C-reactive protein levels were measured in 91 consecutive patients with thrombocytosis (platelet count > or = 600 x 10(9)/L). The cause of thrombocytosis was determined by reviewing the medical histories and follow-up data without knowledge of the corresponding laboratory values. Sixty-four patients had reactive thrombocytosis, 20 had clonal thrombocytosis, and 7 had clonal thrombocytosis plus reactive thrombocytosis. Plasma interleukin-6 was measured by an enzyme-linked immunosorbent assay, and C-reactive protein was measured with rate immunonephelometry. RESULTS: Interleukin-6 levels were undetectable in all the patients with clonal thrombocytosis, whereas they were increased in 60% of the patients with reactive thrombocytosis or clonal thrombocytosis plus reactive thrombocytosis. There was a correlation between interleukin-6 and C-reactive protein levels (r = .6), and the median and range values of both levels differed significantly between the clonal thrombocytosis group and the other two groups (P < 0.0001). In 81% of the patients with reactive thrombocytosis, levels of either interleukin-6 or C-reactive protein were elevated. There was no correlation between interleukin-6 and C-reactive protein levels and the platelet count. CONCLUSIONS: An elevated interleukin-6 level is rare in uncomplicated clonal thrombocytosis and suggests reactive thrombocytosis. However, an isolated normal value has little discriminatory value. Measurement of C-reactive protein level may be used as a less expensive surrogate for measurement of interleukin-6. Repeatedly low levels of both interleukin-6 and C-reactive protein are most consistent with clonal thrombocytosis.

Aged↗