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J Sklar

Publications and source records attributed to J Sklar.

At least 37 records · Page 2Linked to original sources
PubMed1997

Detection of circulating tumor cells in patients with Ewing's sarcoma and peripheral primitive neuroectodermal tumor.

PURPOSE: To determine the feasibility of detecting Ewing's sarcoma (ES) or peripheral primitive neuroectodermal tumor (PNET) through a reverse-transcriptase polymerase chain reaction (RT-PCR) of the t(11;22)(q24;q12) fusion transcript in blood and bone marrow samples from patients with these neoplasms. PATIENTS AND METHODS: Peripheral-blood (PB) and/or bone marrow aspirate (BM) samples were obtained from 28 patients with ES or PNET at initial presentation or at relapse. Patients were divided into two groups: newly diagnosed patients with nonmetastatic disease and those with metastatic/relapsed disease. RNA was extracted from fractionated BM and PB samples, and RT-PCR was performed for the EWS/HumFLI1 fusion mRNA was transcribed across the t(11;22) breakpoint. RESULTS: Among the 16 patients with nonmetastatic disease, three of 16 were RT-PCR positive for EWS/HumFLI1 RNA in BM and three of 10 were positive in PB. The total number of nonmetastatic patients who were positive in either PB or BM was four of 16 (25%). Among patients with metastatic/relapsed disease, two of six were positive in BM and five of 10 were positive in PB. The total fraction of patients with metastatic/relapsed disease that was positive in either BM or PB was six of 12 (50%). CONCLUSION: In this study, we show that it is possible to amplify the EWS/HumFLI1 RNA by RT-PCR from the BM and PB of a subset of patients with both nonmetastatic and metastatic ES or PNET, which implies that occult tumor cells are present at these sites. The true biologic and clinical meaning of this information is unknown. However, it does suggest a possible application of RT-PCR for the monitoring of residual disease in patients who are undergoing therapy for ES or PNET. This approach may permit early identification of patients who may benefit from alternative therapy or who may be spared possible overtreatment.

Chromosomes, Human, Pair 11↗
PubMed1996

Modulated expression of notch1 during thymocyte development.

The Notch gene family encodes transmembrane proteins that have been implicated in control of diverse cellular differentiation events in the fly, frog, and mouse. Mammalian Notch1 is expressed at high levels in thymus and is mutated in a subset of human T-cell acute lymphoblastic neoplasms, suggesting a role in T-cell differentiation. To investigate the patterns of expression of NOTCH1 protein in thymocytes of the developing and mature thymus, antibodies raised against NOTCH1 were used to perform immunohistochemical and flow cytometric analyses. Strong staining for NOTCH1 within the fetal murine thymus was observed as early as 13.5 days postcoitum. By 17.5 days postcoitum, preferential staining of superficial cortical thymocytes was observed, with weak staining of developing medulla. Flow cytometric analysis and immunohistochemical staining of flow-sorted cells confirmed that the highest levels of NOTCH1 expression in adult murine thymus were present in immature cortical thymocytes (CD24high, CD4-CD8-). In contrast, NOTCH1 expression was low or absent in more mature cortical thymocytes (CD24low, CD4+CD8+), whereas intermediate levels of expression were observed in CD4+CD8- and CD4-CD8+ cells. These data indicate a dynamic pattern of NOTCH1 expression during T-cell differentiation and suggest that downregulation of NOTCH1 may be required for maturation of cortical thymocytes.

Animals↗
PubMed1996

Lymphocytic progenitor cell origin and clonal evolution of human B-lineage acute lymphoblastic leukemia.

At presentation, bone marrow specimens from over 25% of B-lineage acute lymphoblastic leukemias (ALL) display more than two clonal rearrangements of immunoglobulin heavy chain (IgH) genes in Southern blot analyses. Nucleotide sequence analysis has shown predominantly different V(H)DJ(H) junctions among these genes, leading to the frequent description of such cases as oligoclonal leukemias. In the present study, we have analyzed the lgH genes from four patients whose leukemic cells contained different patterns of lgH gene rearrangements between presentation and relapse. Nucleotide sequence analysis of the lgH genes showed that three mechanisms could account for these differences: de novo V(H)DJ(H) rearrangement, V(H) to DJ(H) recombination, and V(H) replacement. In all cases, more than two totally different V(H)DJ(H) rearrangements appeared during evolution of the disease, formally consistent with the conclusion that these tumors were composed of apparently unrelated clones. However, the retention of some of the antigen receptor gene rearrangements, as well as the persistence of a chromosomal marker in two cases, indicated that these leukemias had a monoclonal origin. These findings support the hypothesis that some ALLs arise from a lymphoid progenitor cell at a stage of lymphocyte development before the onset of IgH gene rearrangement. These leukemic lymphocyte progenitors generate malignant daughter cells capable of an in vivo maturation that involves the completion of multiple different lgH rearrangements as well as the modification of preexisting rearrangements by V(H) to DJ(H) recombination or by a V(H) replacement.

Base Sequence↗
PubMed1996

Exclusive development of T cell neoplasms in mice transplanted with bone marrow expressing activated Notch alleles.

Notch is a highly conserved transmembrane protein that is involved in cell fate decisions and is found in organisms ranging from Drosophila to humans. A human homologue of Notch, TAN1, was initially identified at the chromosomal breakpoint of a subset of T-cell lymphoblastic leukemias/lymphomas containing a t(7;9) chromosomal translocation; however, its role in oncogenesis has been unclear. Using a bone marrow reconstitution assay with cells containing retrovirally transduced TAN1 alleles, we analyzed the oncogenic potential of both nuclear and extranuclear forms of truncated TAN1 in hematopoietic cells. Although the Moloney leukemia virus long terminal repeat drives expression in most hematopoietic cell types, retroviruses encoding either form of the TAN1 protein induced clonal leukemias of exclusively immature T cell phenotypes in approximately 50% of transplanted animals. All tumors overexpressed truncated TAN1 of the size and subcellular localization predicted from the structure of the gene. These results show that TAN1 is an oncoprotein and suggest that truncation and overexpression are important determinants of transforming activity. Moreover, the murine tumors caused by TAN1 in the bone marrow transplant model are very similar to the TAN1-associated human tumors and suggest that TAN1 may be specifically oncotropic for T cells.

Animals↗
PubMed1996

Loss of heterozygosity at chromosome regions 22q11-12 and 11p15.5 in renal rhabdoid tumors.

Rhabdoid tumors of the kidney are highly malignant neoplasms that occur primarily within the first 3 years of life. Although they are regarded as distinct from Wilms' tumors, their pathogenesis remains unclear. Whereas most cytogenetic studies of these tumors have revealed normal karyotypes, a few reports have indicated abnormalities at chromosome regions 22q and 11p15.5. We analyzed 30 primary renal rhabdoid tumors for loss of heterozygosity (LOH) at both regions and found that 24 of 30 tumors (80%) had LOH at chromosome arm 22q and that 5 of 30 (17%) had LOH at chromosome band 11p15.5. All of the five tumors with LOH at chromosome arm 11p also had LOH at chromosome arm 22q. The data suggest that there is a gene in chromosome 22, probably a tumor suppressor, inactivation of which may be involved in the genesis of renal rhabdoid tumors. A second gene in chromosome segment 11p15.5, in the region of the putative WT2 gene, may also be involved, in at least a subset of rhabdoid tumors. In addition, five tumors were characterized by microsatellite instability at three or more of 21 loci examined, suggesting a possible role for a replicative error in tumorigenesis or progression in some cases of renal rhabdoid tumors. Genes Chromosom Cancer 15:10-17 (1996).

Chromosome Deletion↗
PubMed1996

Hemophagocytosis as a para-neoplastic syndrome in NK cell leukemia.

Hemophagocytic syndrome is a proliferative disorder of an activated monocyte-macrophage system and is characterized by fever, hepato-splenomegaly and pancytopenia. The serum level of interferon-gamma in the syndrome is increased but its origin is unknown. Here we describe a case of NK cell leukemia with hemophagocytic syndrome with elevated serum level of interferon-gamma. The levels of various cytokines were monitored during the course and statistic analysis was performed. To identify the clonal component, the NK cell fraction was sorted from the mononuclear layer and was subjected to Southern blot hybridization with a probe for EB virus tandem repeats. The fraction was also stimulated with interleukin-2 and the level of interferon-gamma in the conditioned medium was measured. Levels of M-CSF and interferon-gamma were significantly correlated with the degree of clinical manifestations and laboratory data. Southern blot hybridization revealed monoclonality of an NK cell fraction. The fraction also released interferon-gamma. Since macrophage can be activated through cytokines, the hemophagocytosis might have been triggered by factor(s) released from the abnormal NK cell clone at least in this case.

Adolescent↗
PubMed1996

Practical methods of mutation detection.

The past few years have seen a marked increase in the demand for practical methods of mutation detection. Over this period of time, both the development of new methods and improvements in existing methods have made mutation detection in the research setting a less cumbersome task. Nevertheless, all of the commonly used approaches to this problem are labor intensive, and truly practical mutation detection methods in the clinical setting will depend on the development of more fully automated technology.

DNA Mutational Analysis↗
PubMed1995

Stress testing with closed-loop arbutamine as an alternative to exercise. The International Arbutamine Study Group.

OBJECTIVES: This study compared exercise and pharmacologic stress testing using arbutamine delivered by a closed-loop device for the detection of coronary artery disease. BACKGROUND: Arbutamine, an agent designed to simulate exercise, has been developed in conjunction with a closed-loop delivery device that modulates the rate of administration on the basis of physiologic feedback. METHODS: Two hundred ten patients (180 men, 30 women) with symptoms and angiographic evidence of coronary artery disease were studied. Ischemia was categorized in three ways: 1) the presence of angina; 2) the occurrence of > or = 0.1-mV horizontal or downsloping ST segment depression or elevation at 60 ms after the J point; or 3) the presence of either condition 1 or 2. RESULTS: In the 210 patients, the mean increase in heart rate and systolic blood pressure evoked by arbutamine and exercise was 51 and 53 beats/min (p = NS) and 36 and 44 mm Hg (p < 0.0001), respectively. Arbutamine detected ischemia more often than exercise with each of the three ischemic end points. Sensitivity for detecting ischemia by either angina or ST segment change was 84% (95% confidence interval ¿ change was 84% (95% confidence interval [CI] 79% to 89%) for arbutamine and 75% (95% CI 69% to 81%) for exercise testing (p = 0.014). For angina alone, sensitivity was 73% (95% CI 67% to 79%) for arbutamine and 64% (95% CI 57% to 71%) for exercise (p = 0.026). For ST segment change alone, sensitivity was 47% (95% CI 40% to 54%) for arbutamine and 44% (95% CI 37% to 51%) for exercise (p = 0.426). Cardiac events occurred in five patients (1.8%) within 24 h of the arbutamine test. CONCLUSIONS: In detecting documented coronary artery disease, the sensitivity of arbutamine testing was equal to that of exercise for the electrocardiographic end point of ST segment change alone. Arbutamine testing was significantly superior to exercise testing for either ST change or angina or for angina alone.

Aged↗
PubMed1995

Frequent loss of heterozygosity at the TEL gene locus in acute lymphoblastic leukemia of childhood.

TEL is a new member of the ETS family of transcription factors which is rearranged in a number of hematologic malignancies with translocations involving chromosome band 12p13. In some cases, both TEL alleles are affected, resulting in loss of wild-type TEL function in the leukemic cells. In addition, 5% of children with acute lymphoblastic leukemia (ALL) have 12p12-p13 deletions, suggesting that a tumor suppressor gene resides on 12p. These observations led us to consider whether TEL loss of function may contribute to the pathogenesis of ALL. In this report we show that the TEL gene maps between the polymorphic markers D12S89 and D12S98, and we use these flanking markers to screen paired diagnosis and remission samples from 81 children with ALL for loss of heterozygosity (LOH) at the TEL gene locus. Fifteen percent of informative patients showed TEL LOH which was not evident on cytogenetic analysis. Detailed examination of patients with LOH at this locus showed that the critically deleted region included two candidate tumor suppressor genes: TEL and KIP1, the gene encoding the cyclin-dependent kinase inhibitor p27. These studies show that LOH at the TEL locus is a frequent finding in childhood ALL.

Adolescent↗
PubMed1995

Gamma delta T cell recognition of tumor Ig peptide.

Although gamma delta T cells have been postulated to act as a surveillance mechanism that eliminates transformed or otherwise damaged cells, little is known about tumor recognition by gamma delta T cells, including the Ags that are recognized and the molecules that present them. Previously, we described human gamma delta CTL that recognize the autologous B cell lymphoma. Here we report that these gamma delta CTL lyse heterologous cells transfected with the tumor Ig lambda chain gene. Furthermore, the lambda chain is recognized as processed peptide in an Id-specific manner. T cell recognition does not involve classical MHC molecules, but it could be blocked by Abs directed against the heat shock protein grp75. These findings show a specific gamma delta T cell response to a highly polymorphic Ag such as tumor Id and implicate heat shock protein as a molecule required for recognition.

Amino Acid Sequence↗
PubMed1995

Lymphomagenesis in the SCID-hu mouse involves abundant production of human interleukin-10.

Both human (hu) and viral (v) interleukin-10 (IL-10) appear to be important cofactors in the survival and growth of lymphoblastoid cell lines infected with Epstein-Barr virus (EBV). When mice with severe combined immune deficiency (SCID) are injected with human peripheral blood lymphocytes (PBL) from normal individuals who are seropositive for EBV, the majority of hu-PBL-SCID mice will develop an EBV-associated lymphoproliferative disease (EBV-LPD) of human B-cell origin, not unlike some cases of EBV-LPD that are seen in immunocompromised individuals. The role of huIL-10 or vIL-10 in this chimeric mouse model of EBV-LPD is unknown. In the present study, we show that hu-PBL-SCID mice that develop EBV-LPD have significant elevation of serum huIL-10 levels compared with mice that do not develop EBV-LPD (P = .005). vIL-10 was undetectable in all animals. The EBV+ tumor samples express transcript for huIL-10 and huIL-10 receptor, express huIL-10 protein by immunohistochemical staining, and show specific binding of recombinant (r) huIL-10. In vitro analysis of the functional consequences of rhuIL-10 binding to IL-10 receptors on fresh EBV+ tumor cells shows that rhuIL-10 can prevent programmed cell death as well as promote proliferation and can do so at concentrations of huIL-10 found in vivo. Thus, huIL-10 production by EBV+ tumor cells may contribute directly to their malignant outgrowth in the hu-PBL-SCID mouse by two autocrine mechanisms: prevention of programmed cell death and proliferation. The implications of such findings with regard to EBV-LPD in humans is discussed.

Animals↗
PubMed1995

Detection of mutations by cleavage of DNA heteroduplexes with bacteriophage resolvases.

We have explored the application of the bacteriophage resolvases T4 endonuclease VII and T7 endonuclease I for detecting mutations in genomic DNA. Heteroduplex DNA fragments prepared by amplification from DNA containing known mutations were cleaved by one or both enzymes at nucleotide mismatches created by 3 of 3 short deletions and 13 of 14 point mutations in fragments as large as 940 basepairs. Heteroduplexes representing all four classes of possible single nucleotide mismatches were cleaved, and the sizes of the cleavage products generated correlated with the location of the mutation. We conclude that bacteriophage resolvases may be useful reagents for the rapid screening of DNA for mutations.

Bacteriophages↗
PubMed1995

Molecular staging of cutaneous T-cell lymphoma: evidence for systemic involvement in early disease.

Biopsies of various tissues from eight patients with confirmed cutaneous T-cell lymphoma were analyzed for lymphomatous involvement using V-J junctional sequences in rearranged T-cell receptor-gamma genes as specific molecular markers for the malignant clone. The patients included one stage IA, one stage IB, and six stage IVA. Twenty-five specimens were analyzed including 14 skin, five lymph node, four blood, and two bone-marrow samples. Ten skin samples and four lymph node samples were histologically positive for lymphoma. The other specimens were morphologically uninvolved. An assay involving polymerase chain reaction (PCR) amplification of T-cell receptor-gamma gene rearrangements and denaturing gradient gel electrophoresis was used to identify the tissue specimen containing the greatest tumor clone density in each case. This specimen was then used to generate a tumor-specific RNA probe that was used to molecularly stage each patient by means of an assay involving PCR gene amplification and RNase protection analysis (PCR/RPA). This assay detected malignant cells in all available biopsies, including morphologically uninvolved extracutaneous tissue samples (two blood, one lymph node, and one bone marrow) obtained from the two patients in pathologic stage I. Microscopic examination and the less sensitive PCR/denaturing gradient gel electrophoresis technique failed to detect lymphomatous involvement in 11 (44%) and eight (32%) of these 25 specimens, respectively. We conclude that molecular biologic staging using PCR/RPA is able to demonstrate morphologically occult dissemination of cutaneous T-cell lymphoma in early disease. In addition, PCR/RPA may be able to monitor tumor response to therapy and detect early recurrence of malignant lymphomas during clinical remission.

Base Sequence↗
PubMed1995

Correlation of loss of heterozygosity at chromosome 9q with histological subtype in medulloblastomas.

Patients with the nevoid basal cell carcinoma syndrome (NBCCS) are at increased risk for medulloblastomas as well as for basal cell carcinomas. The gene for NBCCS has been mapped to chromosome 9q22.3-q31 by linkage analysis, and loss of heterozygosity (LOH) in this region has been demonstrated in approximately one-half of sporadic basal cell carcinomas. In the present study, LOH for chromosome 9q22.3-q31 microsatellite markers was investigated in medulloblastomas occurring among children with NBCCS and in sporadic medulloblastomas. Histologically, all 3 NBCCS medulloblastomas were noted to have a desmoplastic phenotype, and LOH was detected in both of the 2 cases for which microsatellite markers were heterozygous in normal tissues. LOH was also detected in a subset of sporadic medulloblastomas, each of which were found to display the desmoplastic phenotype. In all, 3 of the 6 sporadic desmoplastic tumors showed LOH, whereas LOH was not seen in any of the 11 sporadic, non-desmoplastic medulloblastomas studied. Additionally, desmoplastic tumors lacking detectable LOH each showed histological features of so-called cerebellar neuroblastoma, a subgroup of desmoplastic medulloblastoma with extensive neuroblastomatous differentiation. The data are consistent with a role for inactivation of a tumor suppressor gene at chromosome 9q in the development of medulloblastomas in patients with NBCCS and of sporadic medulloblastomas characterized by a desmoplastic phenotype similar to that found in patients with NBCCS. Restriction of chromosome 9q loss to non-neuroblastomatous desmoplastic tumors suggests that this variant of medulloblastoma maybe pathogenetically distinct from tumors having other histological phenotypes.

Adolescent↗
PubMed1994

Analysis of androgen receptor DNA reveals the independent clonal origins of uterine leiomyomata and the secondary nature of cytogenetic aberrations in the development of leiomyomata.

Uterine leiomyomata are thought to be monoclonal neoplasms. Accordingly, investigations of clonality with G6PD isoforms used as a marker for X chromosome inactivation have suggested independent origins for multiple tumors within individual uteri. However, results from a recent study assessing methylation differences between DNA of active and inactive X chromosomes have been interpreted to suggest that multiple tumors may arise from a common precursor. We have examined the clonality of 36 leiomyomata from 16 patients by analyzing X chromosome inactivation as indicated by the methylation status of the X-linked androgen receptor gene. As shown by this assay, all informative leiomyomata were monoclonal in origin. In patients with multiple leiomyomata, a random distribution of inactivation between the X homologs was noted, consistent with an independent origin of each tumor. Cytogenetic analysis was also performed on short-term cell cultures of 27 of the 36 tumors. In each of two tumors that had both cells with a clonal karyotypic abnormality and karyotypically normal cells, DNA prepared from short-term cultures showed a monoclonal pattern of X inactivation identical to that of the leiomyoma from which they were derived. These data suggest that karyotypically normal cells present in short-term cultures of uterine leiomyomata are part of the tumor clone, and that clonal expansion of tumor cells precedes the development of cytogenetic aberrations.

Alleles↗