Soluble VCAM-1/soluble ICAM-1 ratio is a promising biomarker for diagnosing endometriosis.

Study question: Do cell adhesion molecules play a role in endometriosis, and can they be used as a biomarker for diagnosing endometriosis? Summary answer: Altered expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in the endometrium and peritoneum may play a key role in endometriosis and the soluble VCAM-1/soluble ICAM-1 ratio is a promising biomarker. What is known already: Cell adhesion molecules are cell surface proteins that mediate cellular adherence, inflammatory and immune responses, and cancer-related biological processes. Altered expression of VCAM-1 and ICAM-1 in women with endometriosis has been investigated previously; however, gene expression levels in tissues and protein levels in the serum have not been investigated in the same patients. Study design size, duration: We performed a prospective, longitudinal study (the Endometriosis Marker Austria) in patients who underwent a laparoscopy for benign gynecological pathology in a university-based tertiary referral center for endometriosis. From a total of 138 women who were included in the study from July 2013 through September 2014, 97 had not received hormonal treatment for at least 3 months prior to recruitment and were included in the analysis; 49 (50.5%) of these women had endometriosis, and the 48 (49.5%) who did not have endometriosis served as a control group. Participants/materials setting methods: During laparoscopy, tissue samples were obtained from ectopic and eutopic endometrium, and from normal pelvic peritoneum. In addition, serum samples were collected immediately before and 6-10 weeks after surgery. The mRNA levels of VCAM-1, ICAM-1 and epithelial cell adhesion molecule (EpCAM) were measured using quantitative real-time PCR, and serum protein levels of soluble VCAM-1 (sVCAM-1), ICAM-1 (sICAM-1) and EpCAM (sEpCAM) were measured using ELISA and correlated with endometriosis status. Main results and the role of chance: The mRNA levels of both VCAM-1 and ICAM-1 were higher in ectopic endometriotic lesions than in eutopic endometrium (P < 0.001). Moreover, the mRNA levels of both VCAM-1 and ICAM-1 were higher in normal peritoneum samples obtained from women with endometriosis compared to those from controls (P = 0.038 and P = 0.009). The mRNA levels of VCAM-1 were also higher in the eutopic endometrium samples obtained from women with endometriosis compared to controls (P = 0.018). With respect to serum protein levels, compared to controls, the women with endometriosis had lower serum levels of sICAM-1 (P = 0.042) and higher levels of sVCAM-1 (P < 0.001). Our analysis revealed that the serum levels of sVCAM-1 were not affected by lesion entity, menstrual cycle phase or disease severity. An receiver operating characteristics curve, calculated to determine whether preoperative serum sVCAM-1 concentration can be used to predict endometriosis, found an AUC of 0.868 with 80% specificity and 84% sensitivity at a cutoff value of 370 pg/ml. This predictive performance can be further improved by calculation of the sVCAM-1/sICAM-1 ratio, leading to an AUC of 0.929 with 86.7% specificity and 90.3% sensitivity at a cutoff ratio value of 1.55. Large scale data: Not applicable. Limitations reasons for caution: The relatively small sample size in the expression analyses is a possible limitation of this study. Wider implications of the findings: Our findings could contribute to an improved understanding of the pathogenesis of endometriosis and the role of cell adhesion molecules. In addition, the results may lead to the development of new, non-invasive tools for diagnosing endometriosis. The ability to diagnose patients by measuring serum sVCAM-1 levels or the sVCAM-1/sICAM-1 ratio would have considerable clinical value. Study funding/competing interest(s): The Ingrid Flick Foundation (Grant no. FA751C0801), which played no role in the study design, data collection and analysis, decision to publish or preparation of the manuscript. The authors declare no competing interests.

Next-Generation Sequencing-based genomic profiling of brain metastases of primary ovarian cancer identifies high number of BRCA-mutations.

Ovarian cancer represents the most common gynaecological malignancy and has the highest mortality of all female reproductive cancers. It has a rare predilection to develop brain metastases (BM). In this study, we evaluated the mutational profile of ovarian cancer metastases through Next-Generation Sequencing (NGS) with the aim of identifying potential clinically actionable genetic alterations with options for small molecule targeted therapy. Library preparation was conducted using Illumina TruSight Rapid Capture Kit in combination with a cancer specific enrichment kit covering 94 genes. BRCA-mutations were confirmed by using TruSeq Custom Amplicon Low Input Kit in combination with a custom-designed BRCA gene panel. In our cohort all eight sequenced BM samples exhibited a multitude of variant alterations, each with unique molecular profiles. The 37 identified variants were distributed over 22 cancer-related genes (23.4%). The number of mutated genes per sample ranged from 3 to 7 with a median of 4.5. The most commonly altered genes were BRCA1/2, TP53, and ATM. In total, 7 out of 8 samples revealed either a BRCA1 or a BRCA2 pathogenic mutation. Furthermore, all eight BM samples showed mutations in at least one DNA repair gene. Our NGS study of BM of ovarian carcinoma revealed a significant number of BRCA-mutations beside TP53, ATM and CHEK2 mutations. These findings strongly suggest the implication of BRCA and DNA repair malfunction in ovarian cancer metastasizing to the brain. Based on these findings, pharmacological PARP inhibition could be one potential targeted therapeutic for brain metastatic ovarian cancer patients.

Prenatal microarray analysis as second-tier diagnostic test: single-center prospective study.

OBJECTIVE: To evaluate the usefulness of chromosome microarrays as a second-tier test in prenatal genetic testing. METHODS: We prospectively analyzed 75 high-risk pregnancies undergoing invasive prenatal genetic testing in which the karyotype either was normal or had findings other than a common non-mosaic autosomal aneuploidy. RESULTS: Chromosomal microarray analysis (CMA) was performed successfully in all cases. Pathological copy-number variations (CNVs) explaining the phenotypes were found in 11 cases (14.7%). Four cases were detected with an unbalanced translocation. In three of these cases, subsequent genetic analysis demonstrated that a parent was an unknown carrier of a balanced translocation. Among the 67 cases with normal karyo-types, submicroscopic rearrangements with pathological significance were detected in five (7.5%) and CNVs of unclear significance were detected in one (1.5%). CMA was able to discriminate correctly between true mosaicism and confined or pseudomosaicism in all six mosaic cases. CONCLUSION: CMA is a valuable second-tier test in high-risk pregnancies for which identification or further delineation of genetic aberrations is important. Its higher resolution results in a higher detection rate of aberrant cases, with a clear clinical benefit for estimation of risk of recurrence.

Rituximab monotherapy for primary cutaneous B-cell lymphoma: response and follow-up in 16 patients.

BACKGROUND: We have carried out a retrospective analysis to evaluate the therapeutic value of the anti-CD20 antibody rituximab in 16 consecutive patients with primary cutaneous CD20+ B-cell lymphomas. PATIENTS AND METHODS: Sixteen patients (4 females, 12 males) with a median age of 54 years received systemic therapy with rituximab 375 mg/m(2) once weekly for four or six consecutive weeks. Eleven patients had primary cutaneous follicle center cell lymphoma and five patients had a primary cutaneous marginal zone B-cell lymphoma. RESULTS: Of the 16 patients with PCBCL, 14 patients (87.5%) achieved complete remission (CR). In two patients, partial remission was obtained and additional focal radiotherapy was applied, which resulted in final CR. Five to 14 (35%) patients with CR relapsed, in an interval between 6 and 37 months. There were no severe side-effects. CONCLUSIONS: On the basis of our results, single-agent treatment with i.v. rituximab appears to be feasible and safe and results in a high rate of durable remissions. Judging from our data, it appears to be an attractive treatment option and should be directly compared with local radiotherapy.

Gene expression signature of chronic lymphocytic leukaemia with Trisomy 12.

BACKGROUND: The prognosis of chronic lymphocytic leukaemia (CLL) patients is largely determined by the karyotype of the malignant clone. We have investigated the gene expression profile associated with trisomy 12 (+12). DESIGN: Initially, unselected peripheral blood mononuclear cells of four patients with +12 were compared with 16 CLL controls using microarray analysis. RESULTS: were validated by quantitative real-time PCR with RNA from 61 patients (29 with +12, 32 CLL controls). Results Seven genes showing the strongest correlation with +12 in microarray analysis were selected for real-time PCR: HIP1R, MYF6, SLC2A6, CD9 (overexpressed); CD200, P2RY14, RASGRP3 (underexpressed). Four genes were significantly associated with +12: HIP1R (P<0.0001), MYF6 (P=0.007), P2RY14 (P=0.014), CD200 (P=0.028). Receiver Operating Characteristic curve analysis revealed that HIP1R expression was a highly sensitive and specific marker for +12 in CLL patients. MYF6 was exclusively expressed in normal or malignant B cells in peripheral blood but was poorly predictive for +12. As expected, a number of overexpressed genes are located on chromosome 12 (HIP1R, MYF6). Interestingly, both significantly underexpressed genes (P2RY14, CD200) reside on the long arm of chromosome 3 pointing to trans-repression in this region. CONCLUSIONS: Analysis of the molecular signature of trisomy 12 in CLL resulted in: (i) identification of a surrogate marker for PCR (HIP1R); (ii) observation of a gene dosage effect; and (iii) detection of specific underexpression of genes located on chromosome 3. These results should help to improve diagnosis and treatment decisions for patients with CLL and trisomy 12.

New cellular tools reveal complex epithelial-mesenchymal interactions in hepatocarcinogenesis.

To enable detailed analyses of cell interactions in tumour development, new epithelial and mesenchymal cell lines were established from human hepatocellular carcinoma by spontaneous outgrowth in culture. We obtained several hepatocarcinoma (HCC)-, B-lymphoblastoid (BLC)-, and myofibroblastoid (MF)-lines from seven cases. In-depth characterisation included cell kinetics, genotype, tumourigenicity, expression of cell-type specific markers, and proteome patterns. Many functions of the cells of origin were found to be preserved. We studied the impact of the mesenchymal lines on hepatocarcinogenesis by in vitro assays. BLC- and MF-supernatants strongly increased the DNA replication of premalignant hepatocytes. The stimulation by MF-lines was mainly attributed to HGF secretion. In HCC-cells, MF-supernatant had only minor effects on cell growth but enhanced migration. MF-lines also stimulated neoangiogenesis through vEGF release. BLC-supernatant dramatically induced death of HCC-cells, which could be largely abrogated by preincubating the supernatant with TNFbeta-antiserum. Thus, the new cell lines reveal stage-specific stimulatory and inhibitory interactions between mesenchymal and epithelial tumour cells. In conclusion, the new cell lines provide unique tools to analyse essential components of the complex interplay between the microenvironment and the developing liver cancer, and to identify factors affecting proliferation, migration and death of tumour cells, neoangiogenesis, and outgrowth of additional malignancy.

Chronic autoimmune thyroiditis (Hashimoto's thyroiditis) in patients with MALT lymphoma.

BACKGROUND: Autoimmune diseases have been implicated in the genesis of MALT lymphoma of various localizations. The development of thyroidal MALT lymphoma has been described as an adverse event in patients suffering from long-standing chronic autoimmune thyroiditis (CAT, Hashimoto's thyroiditis). The percentage and possible association between CAT and extrathyroidal MALT lymphoma, however, have not been assessed so far. PATIENTS AND METHODS: A retrospective analysis of 80 patients with MALT lymphoma diagnosed and treated at our institution identified a total of 13 patients (16%) with MALT lymphoma suffering from an underlying CAT. Patient characteristics including site of disease, stage, genetic changes and clinical course were assessed and evaluated. RESULTS: In total, 10 patients were female and 3 male, with the median age being 57 years (range: 31-80). Four patients suffered from thyroidal lymphoma and nine patients had extrathyroidal lymphoma (four gastric, two orbital, one small intestinal and two salivary gland lymphomas). Three patients had a long-standing history of CAT at diagnosis of MALT lymphoma, while CAT was discovered during staging and clinical work-up of MALT lymphoma in the remaining 10 patients. All 13 patients had localized disease, i.e. stage I or II. Only one of the four patients with gastric MALT lymphoma responded to antibiotic treatment against Helicobacter pylori infection. Genetic aberrations were detected in four patients, two of whom had a t(11;18)(q21;q21) translocation, one patient had trisomies 3 and 18 and one had trisomy 18. CONCLUSION: Our findings suggest that CAT is found in patients with not only thyroidal but also nonthyroidal MALT lymphoma. While the nature of our data does not allow for delineation of a direct association between CAT and development of extrathyroidal MALT lymphoma, further prospective studies on this issue are warranted.

Dasatinib inhibits progenitor cell proliferation from polycythaemia vera.

BACKGROUND: A mutation of Janus kinase 2 V617F is present in most patients with polycythaemia vera (PV). However, it is generally believed that JAK2(V617F) is not the sole molecular abnormality in PV. Since dasatinib is currently evaluated in patients with PV, it is of interest to study the effects of dasatinib on the growth of clonal progenitor cells in vitro. DESIGN AND METHODS: Peripheral blood mononuclear cells from patients with PV, chronic myeloid leukaemia (CML) and controls were exposed to dasatinib (0.1 to 500 nm mL(-1)). Colony growth was stimulated by interleukin-3, granulocyte-macrophage colony-stimulating factor and erythropoietin. Endogenous erythroid colony (EEC) growth was investigated without exogenous cytokines. Real-time PCR was performed to assess the percentage of JAK2(V617F) cells. RESULTS: 10 nm of dasatinib suppressed EEC growth from PV by 89% (P = 0.002). This inhibition was dose dependent and occurred at pharmacological concentrations. Erythroid and myeloid colony growth was also significantly suppressed in the presence of exogenous cytokines. When compared to PV the inhibition of stimulated colony growth was significantly less pronounced in controls but tended to be more vigorous in CML. Interestingly, despite the potent inhibition of PV cells real-time PCR revealed that the numbers of JAK2(V617F) transcripts did not decrease upon exposure to dasatinib. CONCLUSION: This study shows a marked inhibition of the proliferative capacity of progenitor cells from PV. Although JAK2(V617F) transcript levels did not decrease upon exposure to dasatinib, the drug might suppress PV progenitors through inhibition of a yet undefined molecular target.

MALT lymphoma in patients with autoimmune diseases: a comparative analysis of characteristics and clinical course.

MALT lymphoma, especially of extragastric origin, is thought to be associated with an underlying autoimmune disease (AD) in a significant proportion of patients. No systematic assessment of the clinical characteristics of MALT lymphoma arising in AD as opposed to patients without AD has been performed so far. Therefore, all patients diagnosed and treated for MALT lymphoma at our institution have prospectively undergone routine clinical and serological assessment for AD since 1997. In total, 158 patients were available for analysis, and 61 out of 158 patients (39%) were diagnosed with an underlying AD. Patients with AD were predominantly women and significantly younger at lymphoma diagnosis (56 versus 67 years, P=0.004), with a significantly higher rate of extragastric lymphomas (P=0.012). Furthermore, lymphomas in these patients showed a lower frequency of trisomy 3 (P=0.04) and a significantly lower response rate to Helicobacter pylori eradication therapy in the case of gastric lymphomas (P=0.03). All other parameters including estimated median time to relapse were comparable between both groups. Our data suggest that patients with AD develop MALT lymphoma significantly earlier in life. The clinical course, however, does not appear to be adversely influenced by the presence of AD, as neither rate of relapse nor times to relapse or survival are significantly different.

Pathology and clinical course of MALT lymphoma with plasmacytic differentiation.

BACKGROUND: The feature of plasmacytic differentiation (PCD) is present in up to 30% of patients diagnosed with mucosa-associated lymphoid tissue (MALT) lymphoma. To date, the influence of PCD on the clinical course of MALT lymphoma has not been assessed. PATIENTS AND METHODS: Therefore, we have retrospectively analysed the clinical characteristics and the course of the disease in 34 (25%) patients with PCD as compared with 101 (75%) MALT lymphoma patients without this histological feature. RESULTS: Patients with PCD had significantly more extragastric lymphomas [28 of 34 (82%) versus 54 of 101 (53%), P = 0.003] and a significantly lower rate of t(11;18) [2 of 26 (8%) versus 22 of 72 (31%), P = 0.02]. There was no significant difference of age at diagnosis (62 versus 64 years, P = 0.64), relapse rate (48% versus 37%, P = 0.27), estimated median time to progression (43 versus 65 months, P = 0.14), monoclonal gammopathy (50% versus 44%, P = 0.63), t(14;18) involving IGH/MALT 1 (11% versus 8%, P = 0.68), trisomy 3 (31% versus 27%, P = 0.69), trisomy 18 (8% versus 10%, P = 0.74) and the presence of autoimmune diseases between both groups (53% versus 37%, P = 0.09). CONCLUSION: In conclusion, we found that PCD is predominantly found in extragastric MALT lymphoma but has no significant impact on clinical course and prognosis.

Mesenchymal stem cells in patients with chronic myelogenous leukaemia or bi-phenotypic Ph+ acute leukaemia are not related to the leukaemic clone.

BACKGROUND: Human mesenchymal stem cells (MSCs) are thought to be multipotent cells which primarily reside in the bone marrow. Besides their well-known ability to replicate as undifferentiated cells and to differentiate into diverse lineages of mesenchymal tissues, they were recently suggested to also give rise to haematopoietic and leukaemic/cancer stem cells. In this study, the relationship between MSCs and leukemic stem cells in patients with either chronic myelogenous leukaemia (CML) or the more primitive variant, Ph+ bi-phenotypic leukaemia was investigated. PATIENTS AND METHODS: Cultured MSCs from 5 patients with CML and 3 patients with bi-phenotypic Ph+ leukaemia, all of them positive for BCP-ABL, were analysed with conventional cytogenetics, fluorescence in situ hybridisation (FISH) and polymerase chain reaction (PCR) for the presence of t(9;22) and BCR-ABL. MSCs were characterised phenotypically with surface markers (+CD73, +CD90, +CD105, -CD34, -CD45) and functionally through their potential to differentiate into both adipocytes and osteoblasts. RESULTS: MSCs could be cultivated from seven patients. These cells were BCR-ABL negative when analysed with conventional cytogenetics and FISH. Further cytogenetic analysis revealed a normal set of chromosomes without any aberrations. Two patients were BCR-ABL-positive when analysed with PCR, probably as a result of MSC contamination with macrophages. CONCLUSION: MSCs in patients with CML or Ph+ bi-phenotypic leukaemia are not related to the malignant cell clone.

Novel t(5;9)(q33;q22) fuses ITK to SYK in unspecified peripheral T-cell lymphoma.

Among peripheral T-cell lymphomas (PTCL), the heterogeneous category of unspecified PTCL represents the most common subtype. Nevertheless, recurrent chromosomal translocations are unknown in this aggressive type of lymphoma. Here we describe a novel t(5;9)(q33;q22) in unspecified PTCL. Molecular analyses delineated the breakpoints to ITK and SYK resulting in a previously undescribed expression of the Syk tyrosine kinase by Itk. ITK-SYK transcripts were detected in five of 30 (17%) unspecified PTCL, but not in cases of angioimmunoblastic T-cell lymphoma (n=9) and anaplastic lymphoma kinase-negative anaplastic large-cell lymphoma (n=7). In all five translocation-positive cases, the breakpoints were identical fusing the N-terminal pleckstrin homology domain and proline-rich region of ITK to the tyrosine kinase domain of SYK. Three of the five t(5;9)(q33;q22)+ unspecified PTCL shared a very similar histological pattern with predominant involvement of lymphoid follicles and the same CD3+CD5+CD4+bcl-6+CD10+ immunophenotype. These results demonstrate the presence of a recurrent t(5;9)(q33;q22) in a subset of unspecified PTCL, which may represent a novel distinct subgroup of PTCL.

T(3;14)(p14.1;q32) involving IGH and FOXP1 is a novel recurrent chromosomal aberration in MALT lymphoma.

The three chromosomal translocations t(11;18)(q21;q21), t(14;18)(q32;q21), and t(1;14)(p22;q32) are associated with MALT lymphoma. In a case of MALT lymphoma of the thyroid, we observed t(3;14)(p14.1;q32) by cytogenetic analysis. Fluorescence in situ hybridization studies showed that the immunoglobulin heavy chain locus (IGH) was rearranged on chromosome 14. Long-distance inverse polymerase chain reaction identified FOXP1 as the partner gene on chromosome 3. To determine the frequency of the t(3;14)(p14.1;q32), two fluorescence in situ hybridization assays were established to screen 91 MALT lymphomas, all of which were negative for the above-mentioned three translocations, and eight splenic and six nodal marginal zone lymphomas. Overall, nine MALT lymphomas (10%) harbored t(3;14)(p14.1;q32) comprising tumors of the thyroid (three of six), ocular adnexa (four of 20), and skin (two of 20), whereas those of the stomach (n = 20), salivary gland (n = 20), and lung (n = 5) were negative as well as the splenic and nodal marginal zone lymphomas. Most t(3;14)(p14.1;q32) + MALT lymphomas harbored additional genetic abnormalities, such as trisomy 3. Further studies revealed that the three known translocations and t(3;14)(p14.1;q32) are mutually exclusive. Real-time quantitative reverse transcriptase polymerase chain reaction showed upregulation of FOXP1 in cases with t(3;14)(p14.1;q32) or trisomy 3. This study identifies FOXP1 as a new translocation partner of IGH in a site-dependent subset of MALT lymphomas.

Variable frequencies of MALT lymphoma-associated genetic aberrations in MALT lymphomas of different sites.

Although several recurrent genetic aberrations are known to occur in MALT lymphoma, no comprehensive study on the most prevalent MALT lymphoma-associated genetic aberrations is available. We therefore screened 252 primary MALT lymphomas for translocations t(11;18)(q21;q21), t(14;18)(q32;q21), and t(1;14)(p22;q32), and trisomies 3 and 18. The above-listed translocations occurred mutually exclusively and were detected overall in 13.5, 10.8, and 1.6% of the cases; trisomy 3 and/or 18 occurred in 42.1%. The frequency at which the translocations occurred varied markedly with the primary site of disease. The t(11;18)(q21;q21) was mainly detected in pulmonary and gastric tumors, whereas the t(14;18)(q32;q21) was most commonly found in lesions of the ocular adnexa/orbit, skin, and salivary glands. Trisomies 3 and 18 each occurred most frequently in intestinal and salivary gland MALT lymphomas. Our results demonstrate that the three translocations and trisomies 3 and 18 occur at markedly variable frequencies in MALT lymphoma of different sites.

Somatic mosaicism of a greater than 1.7-Mb deletion of genomic DNA involving the entire NF1 gene as verified by FISH: further evidence for a contiguous gene syndrome in 17q11.2.

We report on a third case with neurofibromatosis type 1 (NF1) due to mosaicism for a gross deletion in 17q11.2 covering the entire NF1 gene. The deletion was suspected in Giemsa banded chromosomes and was confirmed by fluorescence in situ hybridization using the cosmids CO919 from the 5' region, GO2121 from the central, H10410 from the 3' region of the NF1 gene, and the 1.7-Mb YAC 947G11 spanning the entire 350-kb genomic DNA of the NF1 gene. The deletion was present in 33% of peripheral blood lymphocytes and 58% of fibroblasts. The clinical manifestations in this 6-year-old male patient were especially severe and extended beyond the typical features of NF1. The patient also displayed facial anomalies, severe and early-onset psychomotor retardation, seizures, spasticity, and microcephaly. These features differ from other large-deletion NF1 patients, even nonmosaic cases. The complex phenotype could be explained by the involvement of coding sequences flanking the NF1 gene, thus supporting the existence of a contiguous gene syndrome in 17q11.2.

Familial cryptic translocation with del 4q34-->qter and dup 12pter-->p13 in sibs with tracheal stenosis: clinical, classical and molecular cytogenetic studies and CGH analyses from archival placental tissues evidencing tertiary trisomy 4 in one abortion specimen.

We report on two retarded half-sibs of different sex and seemingly normal karyotype who had the same syndrome of minor anomalies, heart defect and a distal tracheal stenosis, and who shared a healthy mother. These findings raised suspicions of a cryptic chromosome translocation. A translocation t(4;12)(q34;p13), balanced in the mother and unbalanced in the sibs with loss of terminal 4q and gain of terminal 12p regions, was verified by FISH using whole chromosome painting, subtelomeric and YAC probes. Clinical features could be explained by partial monosomy 4q and partial trisomy 12p. Tracheal stenosis was interpreted as a consequence of the same developmental disturbance leading to esophageal atresia and tracheo-esophageal fistula. It was attributed to the 4q deletion in which esophageal atresia as also respiratory difficulties and airway obstructions had been described. Paraffin-embedded placental tissues were available from three of the five abortions of the mother allowing DNA extraction and comparative genome hybridization (CGH). Two of the abortion specimens had the same der(4)t(4;12)(q34;p13) unbalanced translocation as identified in the sibs. In the third abortion specimen, suspicious of triploidy because of partial hydatidiform mole, CGH uncovered a tertiary trisomy 4 resulting from a 3:1 segregation of the translocation chromosomes and their homologs during maternal meiosis I. Differences in CGH results using DNA generated directly or after DOP-PCR were explained by DNA fragmentation in paraffin-embedded tissues and unequal amplification. Am. J. Med. Genet. 94:271-280, 2000.

Amplification of the AML1(CBFA2) gene on ring chromosomes in a patient with acute myeloid leukemia and a constitutional ring chromosome 21.

In the genesis of hematologic neoplasms gene amplification is a mechanism for illegitimate activation of proto-oncogenes. We report a phenotypically normal patient with a constitutional ring chromosome 21 who developed acute myeloid leukemia (AML). The leukemic cells revealed size-variable ring chromosomes 21 with amplification of the proto-oncogene AML1, located in the chromosomal band 21q22, within the rings. Hitherto, amplification of the proto-oncogene AML1-also in form of a ring chromosome-has been described recently only in one patient with myelodysplastic syndrome (MDS). In AML, gene amplification by ring formation has been demonstrated only in another three patients (amplification of the MLL gene in two cases and of the ETV6 gene in one case). Here we present the new evidence that the internal rearrangement of a constitutional ring chromosome 21 resulted in multiplication of a proto-oncogene in bone marrow cells and provided obviously a selective growth advantage. Moreover the amplification of ribosomal DNA was observed in the ring chromosomes of the tumor cells.

Reciprocal translocation (3;5)(q26;q22) and possible BCHE gene involvement in an unusual myelogenous disorder with both myeloproliferative and dysplastic features.

We report on a 77-year-old male patient who presented with an unusual myelogenous disorder exhibiting both myeloproliferative and dysplastic features. The patient suffered from leukocytosis, eosinophilia, basophilia, transfusion dependent anemia, and rapidly progressing thrombocytopenia. Classical chromosome analysis and fluorescence in situ hybridization (FISH) revealed a reciprocal t(3;5)(q26;q22). Using yeast artificial chromosome (YAC) probes, the breakpoint on chromosome 3 was localized to the butyrylcholinesterase (BCHE) gene (3q26.1-q26.2). This gene has recently been implicated in the regulation of myeloid cells. Whether the BCHE gene was also involved in the deregulation of myelopoiesis, causing the unusual clinical picture in this case, remains unknown.

Recruitment of bone-marrow-derived cells by skeletal and cardiac muscle in adult dystrophic mdx mice.

It is commonly accepted, that regenerative capacity of striated muscle is confined to skeletal muscle by activation of satellite cells that normally reside quiescent between the plasmalemma and the basement membrane of muscle fibers. Muscular dystrophies are characterized by repetitive cycles of de- and regeneration of skeletal muscle fibers and by the frequent involvement of the cardiac muscle. Since during the longstanding course of muscular dystrophies there is a permanent demand of myogenic progenitors we hypothesized that this may necessitate a recruitment of additional myogenic precursors from an undifferentiated, permanently renewed cell pool, such as bone marrow (BM) cells. To this end normal and dystrophic (mdx) female mice received bone marrow transplantation (BMT) from normal congenic male donor mice. After 70 days, histological sections of skeletal and cardiac muscle from BMT mice were probed for the donor-derived Y chromosomes. In normal BMT recipients, no Y chromosome-containing myonuclei were detected, either in skeletal or in cardiac muscle. However, in all samples from dystrophic mdx skeletal muscles Y chromosome-specific signals were detected within muscle fiber nuclei, which additionally were found to express the myoregulatory proteins myogenin and myf-5. Moreover, in the hearts of BMT-mdx mice single cardiomyocytes with donor derived nuclei were identified, indicating, that even cardiac muscle cells are able to regenerate by recruitment of circulating BM-derived progenitors. Our findings suggest that further characterization and identification of the BM cells capable of undergoing myogenic differentiation may have an outstanding impact on therapeutic strategies for diseases of skeletal and cardiac muscle.

Differential expression of NF-kappaB target genes in MALT lymphoma with and without chromosome translocation: insights into molecular mechanism.

Mucosa-associated lymphoid tissue (MALT) lymphoma is characterized by t(11;18)(q21;q21)/API2-MALT1, t(1;14)(p22;q32)/BCL10-IGH and t(14;18)(q32;q21)/IGH-MALT1, which commonly activate the nuclear factor (NF)-kappaB pathway. Gastric MALT lymphomas harboring such translocations usually do not respond to Helicobacter pylori eradication, while most of those without translocation can be cured by antibiotics. To understand the molecular mechanism of these different MALT lymphoma subgroups, we performed gene expression profiling analysis of 21 MALT lymphomas (13 translocation-positive, 8 translocation-negative). Gene set enrichment analysis (GSEA) of the NF-kappaB target genes and 4394 additional gene sets covering various cellular pathways, biological processes and molecular functions have shown that translocation-positive MALT lymphomas are characterized by an enhanced expression of NF-kappaB target genes, particularly toll like receptor (TLR)6, chemokine, CC motif, receptor (CCR)2, cluster of differentiation (CD)69 and B-cell CLL/lymphoma (BCL)2, while translocation-negative cases were featured by active inflammatory and immune responses, such as interleukin-8, CD86, CD28 and inducible T-cell costimulator (ICOS). Separate analyses of the genes differentially expressed between translocation-positive and -negative cases and measurement of gene ontology term in these differentially expressed genes by hypergeometric test reinforced the above findings by GSEA. Finally, expression of TLR6, in the presence of TLR2, enhanced both API2-MALT1 and BCL10-mediated NF-kappaB activation in vitro. Our findings provide novel insights into the molecular mechanism of MALT lymphomas with and without translocation, potentially explaining their different clinical behaviors.