High-throughput sequencing of nodal marginal zone lymphomas identifies recurrent BRAF mutations.

Nodal marginal zone lymphoma (NMZL) is a rare small B-cell lymphoma lacking disease-defining phenotype and precise diagnostic markers. To better understand the mutational landscape of NMZL, particularly in comparison to other nodal small B-cell lymphomas, we performed whole-exome sequencing, targeted high-throughput sequencing, and array-comparative genomic hybridization on a retrospective series. Our study identified for the first time recurrent, diagnostically useful, and potentially therapeutically relevant BRAF mutations in NMZL. Sets of somatic mutations that could help to discriminate NMZL from other closely related small B-cell lymphomas were uncovered and tested on unclassifiable small B-cell lymphoma cases, in which clinical, morphological, and phenotypical features were equivocal. Application of targeted gene panel sequencing gave at many occasions valuable clues for more specific classification.

Isolated tumor cells in stage I & II colon cancer patients are associated with significantly worse disease-free and overall survival.

BACKGROUND: Lymph node (LN) involvement represents the strongest prognostic factor in colon cancer patients. The objective of this prospective study was to assess the prognostic impact of isolated tumor cells (ITC, defined as cell deposits ≤ 0.2 mm) in loco-regional LN of stage I & II colon cancer patients. METHODS: Seventy-four stage I & II colon cancer patients were prospectively enrolled in the present study. LN at high risk of harboring ITC were identified via an in vivo sentinel lymph node procedure and analyzed with multilevel sectioning, conventional H&E and immunohistochemical CK-19 staining. The impact of ITC on survival was assessed using Cox regression analyses. RESULTS: Median follow-up was 4.6 years. ITC were detected in locoregional lymph nodes of 23 patients (31.1%). The presence of ITC was associated with a significantly worse disease-free survival (hazard ratio = 4.73, p = 0.005). Similarly, ITC were associated with significantly worse overall survival (hazard ratio = 3.50, p = 0.043). CONCLUSIONS: This study provides compelling evidence that ITC in stage I & II colon cancer patients are associated with significantly worse disease-free and overall survival. Based on these data, the presence of ITC should be classified as a high risk factor in stage I & II colon cancer patients who might benefit from adjuvant chemotherapy.

Molecular staging of lymph node-negative colon carcinomas by one-step nucleic acid amplification (OSNA) results in upstaging of a quarter of patients in a prospective, European, multicentre study.

BACKGROUND: Current histopathological staging procedures in colon carcinomas depend on midline division of the lymph nodes with one section of haematoxylin & eosin (H&E) staining only. By this method, tumour deposits outside this transection line may be missed and could lead to understaging of a high-risk group of stage UICC II cases, which recurs in ∼20% of cases. A new diagnostic semiautomated system, one-step nucleic acid amplification (OSNA), detects cytokeratin (CK) 19 mRNA in lymph node metastases and enables the investigation of the whole lymph node. The objective of this study was to assess whether histopathological pN0 patients can be upstaged to stage UICC III by OSNA. METHODS: Lymph nodes from patients who were classified as lymph node negative after standard histopathology (single (H&E) slice) were subjected to OSNA. A result revealing a CK19 mRNA copy number >250, which makes sure to detect mainly macrometastases and not isolated tumour cells (ITC) or micrometastases only, was regarded as positive for lymph node metastases based on previous threshold investigations. RESULTS: In total, 1594 pN0 lymph nodes from 103 colon carcinomas (median number of lymph nodes per patient: 14, range: 1-46) were analysed with OSNA. Out of 103 pN0 patients, 26 had OSNA-positive lymph nodes, resulting in an upstaging rate of 25.2%. Among these were 6/37 (16.2%) stage UICC I and 20/66 (30.3%) stage UICC II patients. Overall, 38 lymph nodes were OSNA positive: 19 patients had one, 3 had two, 3 had three, and 1 patient had four OSNA-positive lymph nodes. CONCLUSIONS: OSNA resulted in an upstaging of over 25% of initially histopathologically lymph node-negative patients. OSNA is a standardised, observer-independent technique, allowing the analysis of the whole lymph node. Therefore, sampling bias due to missing investigation of certain lymph node tissue can be avoided, which may lead to a more accurate staging.

Surface atom motion to move iron nanocrystals through constrictions in carbon nanotubes under the action of an electric current.

Under the application of electrical currents, metal nanocrystals inside carbon nanotubes can be bodily transported. We examine experimentally and theoretically how an iron nanocrystal can pass through a constriction in the carbon nanotube with a smaller cross-sectional area than the nanocrystal itself. Remarkably, through in situ transmission electron imaging and diffraction, we find that, while passing through a constriction, the nanocrystal remains largely solid and crystalline and the carbon nanotube is unaffected. We account for this behavior by a pattern of iron atom motion and rearrangement on the surface of the nanocrystal. The nanocrystal motion can be described with a model whose parameters are nearly independent of the nanocrystal length, area, temperature, and electromigration force magnitude. We predict that metal nanocrystals can move through complex geometries and constrictions, with implications for both nanomechanics and tunable synthesis of metal nanoparticles.

Prognostic and predictive value of TOPK stratified by KRAS and BRAF gene alterations in sporadic, hereditary and metastatic colorectal cancer patients.

BACKGROUND: Our aim was to investigate the prognostic and predictive value of the oncogenic MAPKK-like protein T-cell-originated protein kinase (TOPK) stratified by KRAS and BRAF mutations in patients with sporadic, hereditary and metastatic colorectal cancer (CRC) treated with anti-EGFR therapy. METHODS: Immunohistochemistry (IHC) for TOPK was performed on four study groups. Group 1 included two subgroups of 543 and 501 sporadic CRC patients used to test the reliability of TOPK expression by IHC. In Group 2, representing an additional 222 sporadic CRCs, the prognostic effect of TOPK stratified by KRAS and BRAF was assessed. The prognostic effect of TOPK was further analysed in Group 3, representing 71 hereditary Lynch syndrome-associated CRC patients. In Group 4, the predictive and prognostic value of TOPK was analysed on 45 metastatic patients treated with cetuximab or panitumumab stratified by KRAS and BRAF gene status. RESULTS: In both sporadic CRC subgroups (Group 1), associations of diffuse TOPK expression with clinicopathological features were reproducible. Molecular analysis of sporadic CRCs in Group 2 showed that diffuse TOPK expression was associated with KRAS and BRAF mutations (p<0.001) and with poor outcome in patients with either mutation in univariate and multivariate analysis (P=0.017). In hereditary patients (Group 3), diffuse TOPK was linked to advanced pT stage. In metastatic patients treated with anti-EGFR therapy (Group 4), diffuse TOPK expression was linked to dismal outcome despite objective response to treatment (P=0.01). CONCLUSIONS: TOPK expression is an unfavourable prognostic indicator in sporadic patients with KRAS or BRAF mutations and also in patients with metastatic disease experiencing a response to anti-EGFR therapies. The inhibition of TOPK, which could benefit 30-40% of CRC patients, may represent a new avenue of investigation for targeted therapy.

[Lymphomas of the spleen]. / Lymphome der Milz.

The spleen is commonly affected by malignant lymphomas and the macroscopic findings of the spleen correlate with different lymphoma entities. However, most lymphomas are not primarily diagnosed in splenectomy specimens. Exceptions include splenic marginal zone lymphomas and hepatosplenic T-cell lymphomas that are typically diagnosed from histological findings. In addition, hairy-cell leukemia, LGL leukemia and T-cell prolymphocytic leukemia typically show characteristic patterns of infiltration in the spleen which may be diagnostically useful. The different infiltration patterns of these tumors are discussed here.

[Functional splenic pathology and differential diagnosis in splenectomy]. / Funktionelle Pathologie der Milz. Grundlagen und differenzialdiagnostische Erwägungen am Milzpräparat.

From a pathologic-anatomic perspective, the splenic parenchyma is divided into red and white pulp, which react to a certain extent independently from one another in the context of systemic and local processes. While the red pulp serves to filter blood, the white pulp, together with the perifollicular marginal zone, organizes immune reactions. In the marginal zone, innate and acquired immune responses are integrated. The vast majority of splenectomies serve therapeutic purposes in the context of already known disease. Therefore, only about 10% of splenectomy specimens are diagnostic challenges, including the diagnosis of haematological diseases, splenomegaly or splenic tumors. Differential diagnosis considerations are discussed based on the clinical presentation.

Amplification at 7q22 targets cyclin-dependent kinase 6 in T-cell lymphoma.

Recurrent chromosomal aberrations in hematopoietic tumors target genes involved in pathogenesis. Their identification and functional characterization are therefore important for the establishment of rational therapies. Here, we investigated genomic amplification at 7q22 in the T-cell lymphoma cell line SU-DHL-1 belonging to the subtype of anaplastic large-cell lymphoma (ALCL). Cytogenetic analysis mapped this amplicon to 86-95 Mb. Copy-number determination quantified the amplification level at 5- to 6-fold. Expression analysis of genes located within this region identified cyclin-dependent kinase 6 (CDK6) as a potential amplification target. In comparison with control cell lines, SU-DHL-1 expressed considerably higher levels of CDK6. Functionally, SU-DHL-1 cells exhibited reduced sensitivity to rapamycin treatment, as indicated by cell growth and cell cycle analysis. Rapamycin reportedly inhibits degradation of the CDK inhibitor p27 with concomitant downregulation of cyclin D3, implying a proliferative advantage for CDK6 overexpression. Amplification of the CDK6 locus was analyzed in primary T-cell lymphoma samples and, while detected infrequently in those classified as ALCL (1%), was detected in 23% of peripheral T-cell lymphomas not otherwise specified. Taken together, analysis of the 7q22 amplicon identified CDK6 as an important cell cycle regulator in T-cell lymphomas, representing a novel potential target for rational therapy.

Nanotube radio.

We have constructed a fully functional, fully integrated radio receiver from a single carbon nanotube. The nanotube serves simultaneously as all essential components of a radio: antenna, tunable band-pass filter, amplifier, and demodulator. A direct current voltage source, as supplied by a battery, powers the radio. Using carrier waves in the commercially relevant 40-400 MHz range and both frequency and amplitude modulation techniques, we demonstrate successful music and voice reception.

[Phlegmonous gastritis in a 44-year-old woman on immunosuppressive therapy]. / Phlegmonöse Gastritis bei einer 44-jährigen Patientin unter immunsuppressiver Therapie.

HISTORY AND ADMISSION FINDINGS: A 44-year-old woman was on long-term immunosuppressive therapy with leflunomide and adalimumab for rheumatoid arthritis. She was admitted to the emergency room with diffuse abdominal pain of sudden onset. On physical examination she had rebound tenderness in all four abdominal quadrants. INVESTIGATIONS: The white blood cell count was 3300/l, C-reactive protein 25 mg/dl and serum lactate 10 mmol/l. Abdominal computed tomography revealed a diffusely thickened gastric wall and ascites. At explorative laparotomy 1000 ml of a cloudy peritoneal fluid were aspirated and found to be negative for bacteria. But a culture of a mesenterial smear grew streptococci group A. Intra-operative endoscopy showed extensive hemorrhagic gastritis. Because there was no perforation or transmural necrosis gastric resection was not performed. DIAGNOSIS, TREATMENT AND COURSE: Diffuse thickening of the gastric wall, extended mucosal necrosis and the peritoneal finding of streptococci in an immunocompromised patient suggested the diagnosis of phlegmonous gastritis. On treatment with antibiotics and proton pump inhibitor the patient made a slow recovery over the following eight weeks. Nine months after the event an asymptomatic antral stricture was noticed at follow-up gastroscopy. CONCLUSION: Phlegmonous gastritis is a rare but life-threatening complication in immunosuppressed patients.

Tissue microarray-based screening for chromosomal breakpoints affecting the T-cell receptor gene loci in mature T-cell lymphomas.

The pathogenesis of mature T-cell non-Hodgkin lymphomas (T-NHLs) is poorly understood. Analogous to B-cell lymphomas, in which the immunoglobulin (IgH) receptor loci are frequently targeted by chromosomal translocations, the T-cell receptor (TCR) gene loci are affected by translocations in a subset of precursor T-cell malignancies. In a large-scale analysis of 245 paraffin-embedded mature T-NHLs, arranged in a tissue microarray format and using improved FISH assays for the detection of breakpoints in the TCRalpha/delta, TCRbeta, and TCRgamma loci, we provide evidence that mature T-NHLs other than T-cell prolymphocytic leukaemia (T-PLL) also occasionally show a chromosomal rearrangement that involves the TCRalpha/delta locus. In particular, one peripheral T-cell lymphoma (not otherwise specified, NOS) with the morphological variant of Lennert lymphoma displayed a chromosomal translocation t(14;19) involving the TCRalpha/delta and the BCL3 loci. A second case, an angio-immunoblastic T-cell lymphoma (AILT), carried an inv(14)(q11q32) affecting the TCRalpha/delta and IgH loci. FISH signal constellations as well as concomitant comparative genomic hybridization (CGH) data were also suggestive of the occurrence of an isochromosome 7, previously described to be pathognomonic for hepatosplenic T-cell lymphomas, in rare cases of enteropathy-type T-cell lymphoma.

[Enteropathy type T-cell lymphomas: pathology and pathogenesis]. / Pathologie und Pathogenese des Enteropathie-assoziierten T-Zell-Lymphoms.

Enteropathy type T-cell lymphomas (ETL) are the most frequent T-cell lymphomas arising in the gastrointestinal tract. Commonly, the neoplasm is clinically associated with symptoms of malabsorption, and it frequently manifests as a spontaneous bowel perforation. Among ETL, two types can be distinguished by morphology, immunophenotype and, possibly, by pathogenesis. A total of 80% of ETL are characterized by a close association with celiac disease, pleomorphic cytomorphology and the rare expression of CD8 and CD56. In contrast, 20% of ETL show a monomorphic small to medium size cytomorphology and frequent expression of CD8 and CD56, an association with celiac disease is rare in the latter cases. Genetically, ETL is characterized by frequent and recurrent chromosomal gains of 9q33-q34.

[Peripheral NK/T-cell lymphoma]. / Periphere NK/T-Zell-Lymphome.

Peripheral T-cell lymphomas comprise 8% of the malignant lymphomas in Germany. About 25% of these cases present primarily in extranodal localizations. Such localizations are typical for the respective disease and form the basis for the classification of extranodal peripheral T-cell lymphoma. The morphology, immunophenotype and lineage specificity of the tumor cells (originating from T- or NK-cells) is only secondary for the classification. Extranodal NK/T-cell lymphomas of the nasal type are characterized by an angiocentric growth pattern and large confluent areas of necrosis. In addition, there is a clonal infection by Epstein-Barr virus in the T-lymphocytes. In the differential diagnosis, B-cell lymphomas are more frequent at all localizations than T- or NK-cell lymphomas.

Corpuscular thymoma: entity or variant of organotypical thymomas WHO B2/B3?

Hassall's corpuscles are regular structures in the medulla of the normal thymus and in non-neoplastic thymic conditions, e.g. in multilocular thymic cysts. In thymomas, however, they are inconsistently found, and are believed to indicate medullary differentiation of WHO type B1-3 thymomas. We present five organotypical thymomas resembling WHO type B2 and B3 thymomas, but with an abundance of Hassall's corpuscles. We wonder whether this exceedingly rare observation might herald a distinct entity. Four tumors were asymptomatic, incidental findings and of low Masaoka stage (I or II) [20] . One patient suffered from myasthenia gravis which disappeared upon surgical removal of the thymus, while all other patients had no concomitant autoimmune disease. Two patients had a relapse-free follow-up of 12 and 2 years, respectively, upon curative surgery, and another tumor was an autopsy finding; follow-up data of two more recent cases was not yet available. The neoplastic epithelium other than Hassall's corpuscles was arranged either in a cortical type B2 pattern or in type B3 solid cords. In all examples, there was cyst formation, inflammatory reaction and repair, indicative of a long-standing condition. Immature T cells were present in all instances. "Corpuscular thymomas" morphologically resemble WHO type B2 and B3 thymomas, but appear biologically indolent and are rarely associated with myasthenia gravis. Whether they qualify for a separate entity has to be proven by larger series, including genetic studies.

Composite marginal zone B-cell lymphoma and classical Hodgkin's lymphoma: a clinicopathological study of 12 cases.

AIMS: Classical Hodgkin's lymphoma (cHL) rarely coexists as composite lymphoma with B-cell non-Hodgkin's lymphoma (B-NHL). We characterized 12 cases of composite marginal zone B-cell lymphoma (MZBL) and cHL by immunohistochemistry and molecular biology. METHODS AND RESULTS: Eight patients had gastric MZBL of mucosa-associated lymphoid tissue (MALT)-type, in five cases with a diffuse large B-cell lymphoma component. Concurrent cHL was observed either in the stomach wall, regional, or distant lymph nodes. One patient each had composite pulmonary/thyroid MZBL of MALT-type and cHL. In two cases, nodal composite MZBL and cHL was observed. cHL displayed features of mixed cellularity type in 10 cases, while in two cases only scattered Hodgkin- and Reed-Sternberg (H/RS) cells were noted. H/RS cells expressed CD30, multiple myeloma oncogene 1 protein (MUM1P), p53 (100%), CD15 (58%), CD20 (58%) and Epstein-Barr virus-associated LMP1 (50%). No t(11;18)(q21;q21) was detected in composite MZBL of MALT-type and cHL. CONCLUSIONS: MZBL and cHL may occur as composite lymphoma, possibly reflecting clonal lymphoma progression. Derivation from extranodal MZBL of MALT-type should be excluded in cases in which a diagnosis of primary extranodal cHL is considered.

[Anaplastic large cell lymphomas lack the expression of T-cell receptor molecules]. / Fehlende T-zell Rezeptor Expression in grosszellig anaplastischen T-Zell Lymphomen.

Anaplastic large cell lymphoma (ALCL) designates a heterogeneous group of CD30+ (systemic or primary cutaneous) peripheral T-cell lymphomas (PTCLs). A subgroup of systemic ALCL is transformed by anaplastic lymphoma kinase (ALK). We compared 46 ALCL with 22 PTCLs in terms of T-cell receptor (TCR) rearrangements, expression of TCRs and TCR-associated molecules [CD3, ZAP-70 (zeta-associated protein 70)]. Despite their frequent clonal rearrangement for TCRbeta, only 4% of ALCLs expressed TCRbeta protein, whereas TCRs were detected in 86% of PTCLs. Moreover, both TCRbeta+ ALCLs lacked CD3 and ZAP-70 (ie, molecules indispensable for the transduction of cognate TCR signals). Defective expression of TCRs is a common characteristic of all types of ALCL, which may contribute to the dysregulation of intracellular signaling pathways controlling T-cell activation and survival. This molecular hallmark of ALCL is analogous to defective immunoglobulin expression distinguishing Hodgkin lymphoma from other B-cell lymphomas.

What is CD4+CD56+ malignancy and how should it be treated?

CD4+CD56+ malignancy is a rare neoplasm with a typical clinical pattern, an aggressive course and high early relapse rate despite good initial response to chemotherapy. In this review, the impact of different therapeutic approaches on clinical outcome has been studied. We evaluated 91 published cases and our own six patients in terms of clinical features, immunophenotype/cytogenetics and treatment outcome. Treatment was divided into four groups: (A) chemotherapy less intensive than CHOP; (B) CHOP and CHOP-like regimens; (C) therapy for acute leukemia; (D) allogeneic/autologous stem cell transplantation. The median overall survival was only 13 months for all patients. Patients with skin-restricted disease showed no difference in the overall survival from patients with advanced disease (17 and 12 months, respectively). Age >/=60 years was a negative prognostic factor. Age-adjusted analysis revealed improved survival after high-dose chemo/radiotherapy followed by allogeneic stem cell transplantation when performed in first complete remission. This therapeutic approach should be recommended for eligible patients with CD4+CD56+ malignancy. For older patients the best treatment option is still unknown.

Immunohistochemical analysis of B-cell lymphoma using tissue microarrays identifies particular phenotypic profiles of B-cell lymphomas.

AIMS: To validate the applicability of tissue microarray (TM) in immunohistochemical profiling of B-cell lymphoma and to identify particular phenotypic profiles of B-cell neoplasms. METHODS AND RESULTS: Eighty-two diffuse large B-cell lymphomas (DLBL), 54 follicular lymphomas (FL) and 74 mantle cell lymphomas (MCL) were arrayed. Immunohistochemical stains of TM were compared with immunostains of conventional, formalin-fixed and frozen material sections. Concordant staining results were obtained in more than 88% of cases for CD20, CD3, CD5, CD10, CD23, Bcl-2, IgD, secretory differentiation, p53 and p21 expression. Prognostically relevant hot-spot expression of Ki67 yielded concordant results in 71%. Applying TM for characterization of p27KIP1 expression, both typical and blastoid MCL only rarely showed p27KIP1 expression (9% and 15%), whereas 32% of nodal DLBL were p27KIP1-positive, irrespective of high proliferative activity. Among 22 B-cell lymphomas investigated genetically, a p53 + p21- immunophenotype in >20% of tumour cells correlated with p53 locus deletion. CONCLUSIONS: Lymphoma TM allows for immunohistochemical profiling of human B-cell lymphoma with a comparable accuracy to immunohistochemical studies performed on conventional tissue sections. Nodal DLBLs showed significantly more frequent expression of IgD and p27KIP1 than extranodal DLBL. MCL and DLBL frequently showed aberrant p27KIP1 expression. A p53 + p21- immunophenotype in >20% of tumour cells in B-cell non-Hodgkin's lymphoma correlates with p53 gene deletion.