Variation in pre-PCR processing of FFPE samples leads to discrepancies in BRAF and EGFR mutation detection: a diagnostic RING trial.

Aims: Mutation detection accuracy has been described extensively; however, it is surprising that pre-PCR processing of formalin-fixed paraffin-embedded (FFPE) samples has not been systematically assessed in clinical context. We designed a RING trial to (i) investigate pre-PCR variability, (ii) correlate pre-PCR variation with EGFR/BRAF mutation testing accuracy and (iii) investigate causes for observed variation.

Methods: 13 molecular pathology laboratories were recruited.

EuroClonality/BIOMED-2 guidelines for interpretation and reporting of Ig/TCR clonality testing in suspected lymphoproliferations.

PCR-based immunoglobulin (Ig)/T-cell receptor (TCR) clonality testing in suspected lymphoproliferations has largely been standardized and has consequently become technically feasible in a routine diagnostic setting. Standardization of the pre-analytical and post-analytical phases is now essential to prevent misinterpretation and incorrect conclusions derived from clonality data. As clonality testing is not a quantitative assay, but rather concerns recognition of molecular patterns, guidelines for reliable interpretation and reporting are mandatory.

Serous fluid cytology of multicentric Castleman's disease and other lymphoproliferative disorders associated with Kaposi sarcoma-associated herpes virus: a review with case reports.

Objective: The aim of this study is to describe and review the cytological features of Kaposi sarcoma-associated herpes virus (KSHV) related entities, such as multicentric Castleman's disease (MCD), plasmablastic-lymphoma (PBL) and primary effusion lymphoma (PEL), which all may present as body cavity effusions. Serous fluid cytology of MCD and PBL has not, to our knowledge, thus far been described. Although different in nature, MCD, PBL and PEL are characterized by similar morphological features.

PCR-based tissue identification: the UCLH experience.

The need to accurately identify tissue of an individual can arise in a variety of settings including mislabelled slides or sample carryover. Reported rates of carryover range from 0.6% to 2.

IDH1 and IDH2 mutations are frequent events in central chondrosarcoma and central and periosteal chondromas but not in other mesenchymal tumours.

Somatic mutations in isocitrate dehydrogenase 1 (IDH1) and IDH2 occur in gliomas and acute myeloid leukaemia (AML). Since patients with multiple enchondromas have occasionally been reported to have these conditions, we hypothesized that the same mutations would occur in cartilaginous neoplasms. Approximately 1200 mesenchymal tumours, including 220 cartilaginous tumours, 222 osteosarcomas and another ∼750 bone and soft tissue tumours, were screened for IDH1 R132 mutations, using Sequenom(®) mass spectrometry.

Polymerase chain reaction-based clonality analysis of cutaneous B-cell lymphoproliferative processes.

Introduction: The differential diagnosis of B-cell lymphoproliferative processes remains a challenge for pathologists, dermatologists and oncologists, despite advances in histology, immunohistochemistry and molecular biology.

Objective: Evaluate aid and limitations of clonality analysis in the diagnosis of primary cutaneous B-cell lymphomas and B-cell pseudolymphomas.

Methods: This study included 29 cases of B-cell lymphoproliferative processes classified as primary cutaneous B-cell lymphomas (13), B-cell pseudolymphomas (6) and inconclusive cases (10) using histology and immunohistochemistry.

Familial tumoral calcinosis and hyperostosis-hyperphosphataemia syndrome are different manifestations of the same disease: novel missense mutations in GALNT3.

Objective: To report on the biochemistry and clinical and genetic findings of two siblings, the younger sister presenting with recurrent bone pain of the radius and ulna, and medullary sclerosis, and the older brother with soft tissue calcific deposits (tumoral calcinosis) but who later developed bone pain. Both were found to be hyperphosphaturic.

Materials And Methods: The index family comprised four individuals (father, mother, brother, sister).

Clear cell sarcoma of the mediastinum.

A 59-year-old woman presented with a large mediastinal mass. At thoracotomy, the mass was found tightly adherent to the esophageal wall and right lower lobe of the lung. Histological examination showed a solid tumor composed of closely packed nests of cells with clear and eosinophilic cytoplasm, which were strongly and diffusely positive for S100 protein but negative for HMB45 and Melan-A.

Analysis of the fibroblastic growth factor receptor-RAS/RAF/MEK/ERK-ETS2/brachyury signalling pathway in chordomas.

Chordomas are rare primary malignant bone tumours that derive from notochord precursor cells and express brachyury, a molecule involved in notochord development. Little is known about the genetic events responsible for driving the growth of this tumour, but it is well established that brachyury is regulated through fibroblastic growth factor receptors (FGFRs) through RAS/RAF/MEK/ERK and ETS2 in ascidian, Xenopus and zebrafish, although little is known about its regulation in mammals. The aim of this study was to attempt to identify the molecular genetic events that are responsible for the pathogenesis of chordomas with particular focus on the FGFR signalling pathway on the basis of the evidence in the ascidian and Xenopus models that the expression of brachyury requires the activation of this pathway.

Potential therapeutic targets for chordoma: PI3K/AKT/TSC1/TSC2/mTOR pathway.

Chordomas are radio- and chemo-resistant tumours and metastasise in as many as 40% of patients. The aim of this study was to identify potential molecular targets for the treatment of chordoma. In view of the reported association of chordoma and tuberous sclerosis complex syndrome, and the available therapeutic agents against molecules in the PI3K/AKT/TSC1/TSC2/mTOR pathway, a tissue microarray of 50 chordoma cases was analysed for expression of active molecules involved in this signalling pathway by immunohistochemistry and a selected number by western blot analysis.

GNAS1 mutations occur more commonly than previously thought in intramuscular myxoma.

Mutation detection plays an important role in diagnostic pathology, not only in providing a tissue diagnosis, but also in predicting response to antitumourigenic agents. However, mutation detection strategies are often hampered by masking of mutant alleles by wild-type sequences. Coamplification at lower denaturation temperature PCR (COLD-PCR) reportedly increases the proportion of rare variant sequences in a wild-type background by using PCR cycles in which the denaturation temperature is reduced to favour product formation with lower melt temperatures and heteroduplexes arising from minor variants.

Follicular lymphoma of the thyroid gland.

The majority of lymphomas arising in the thyroid gland are mucosa-associated lymphoid tissue lymphomas and diffuse large B-cell lymphomas, which arise from a background of chronic lymphocytic thyroiditis. Follicular lymphoma may also present in the thyroid gland, but its clinicopathologic features at this site are not well characterized, leading to difficulties in diagnosis and clinical management. We have addressed this problem by studying the clinical, morphologic, immunophenotypic, and genetic features of 22 such cases.

Detection of beta-catenin mutations in paraffin-embedded sporadic desmoid-type fibromatosis by mutation-specific restriction enzyme digestion (MSRED): an ancillary diagnostic tool.

Desmoid-type fibromatosis is a locally aggressive deep soft tissue tumor. Some cases are associated with adenosis polyposis coli germline mutations whereas others harbor somatic beta-catenin point mutations mainly in exon 3, codons 41 and 45. These mutations result in stabilization of beta-catenin, and activation of the Wnt signaling pathway.

Histologic evolution of angioimmunoblastic T-cell lymphoma in consecutive biopsies: clinical correlation and insights into natural history and disease progression.

Angioimmunoblastic T-cell lymphoma (AITL) is an uncommon, but aggressive nodal peripheral T-cell lymphoma. Little is known of its biology and its natural history has been poorly studied. We report the first comprehensive study on the natural history/histologic progression of AITL by reviewing consecutive biopsies in 31 cases.

Primary follicular lymphoma of the testis and epididymis in adults.

Primary testicular lymphomas typically occur in patients over 60 years of age. Most are diffuse large B-cell lymphomas with frequent dissemination and a poor prognosis. Primary follicular lymphoma of the adult testis has not been well characterized.

Angioimmunoblastic T-cell lymphoma: histological progression associates with EBV and HHV6B viral load.

The clinical and histological presentations of angioimmunoblastic T-cell lymphoma (AITL) often mimic an infectious process. Epstein-Barr virus (EBV) and human herpes virus (HHV6) are known to be associated with AITL, but whether these viral infections play a role in its pathogenesis is unclear. It also remains to be investigated whether there might be other viruses associated with AITL.

Mutations in SH3BP2, the cherubism gene, were not detected in central or peripheral giant cell tumours of the jaw.

Giant cell granulomas of the jaw (GCGJ) are non-familial, generally unilateral osteoclast-rich lesions that are histopathologically indistinguishable from cherubism. Cherubism is an autosomal dominant disease that is characterised by bilateral radiolucencies of the jaw, and caused by mutations that occur in SH3BP2 exon 10. The aim of the study was to screen lesional GCGJ tissue for SH3BP2 mutations.

A sensitive mutation-specific screening technique for GNAS1 mutations in cases of fibrous dysplasia: the first report of a codon 227 mutation in bone.

Aims: To report on the mutation-specific restriction enzyme digest (MSRED) method using paraffin-embedded tissue as a means of detecting GNAS1 mutations in fibrous dysplasia (FD), and to determine if any of the reported GNAS1 mutations in endocrine neoplasms, not previously documented in FD, can be found in FD.

Methods And Results: Sixty-seven cases of extragnathic FD were analysed as two groups, 1997-2002 and 2003-06, chosen because tissue fixation and decalcification methods were more accurately recorded in the latter. MSRED revealed that between 2003 and 2006, 93% of 28 'in house' extragnathic cases harboured a GNAS1 mutation, compared with 75% of 32 cases before 2003.

Distinguishing angioimmunoblastic T-cell lymphoma from peripheral T-cell lymphoma, unspecified, using morphology, immunophenotype and molecular genetics.

Aims: To identify distinguishing histological, immunophenotypic and molecular genetic features between angioimmunoblastic T-cell lymphoma (AITL) and peripheral T-cell lymphoma (PTL).

Methods: Nodal T-cell lymphomas examined (n =137), included AITL (n = 89), PTL (n = 22), anaplastic large cell lymphoma (n = 16) and 'AITL/PTL indeterminate' (n = 10) with overlapping features between AITL and PTL, showing morphology typical of AITL but lacking follicular dendritic cell expansion. Immunohistochemistry for CD3, CD20, CD21 and CD10, in situ hybridization for Epstein-Barr virus encoded RNA (EBER) and polymerase chain reaction for T-cell and B-cell clonality analysis were performed.

Detection of SS18-SSX fusion transcripts in formalin-fixed paraffin-embedded neoplasms: analysis of conventional RT-PCR, qRT-PCR and dual color FISH as diagnostic tools for synovial sarcoma.

Synovial Sarcoma consistently harbors t(X;18) resulting in SS18-SSX1, SS18-SSX2 and rarely SS18-SSX4 fusion transcripts. Of 328 cases included in our study, synovial sarcoma was either the primary diagnosis or was very high in the differential diagnosis in 134 cases: of these, amplifiable cDNA was obtained from 131. SS18-SSX fusion products were found in 126 (96%) cases (74 SS18-SSX1, 52 SS18-SSX2), using quantitative and 120 by conventional reverse transcriptase-polymerase chain reaction (RT-PCR).

Significantly improved PCR-based clonality testing in B-cell malignancies by use of multiple immunoglobulin gene targets. Report of the BIOMED-2 Concerted Action BHM4-CT98-3936.

Polymerase chain reaction (PCR) assessment of clonal immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements is an important diagnostic tool in mature B-cell neoplasms. However, lack of standardized PCR protocols resulting in a high level of false negativity has hampered comparability of data in previous clonality studies. In order to address these problems, 22 European laboratories investigated the Ig/TCR rearrangement patterns as well as t(14;18) and t(11;14) translocations of 369 B-cell malignancies belonging to five WHO-defined entities using the standardized BIOMED-2 multiplex PCR tubes accompanied by international pathology panel review.