Publications by authors named "Alexandra Clipson"

15 Publications

  • Page 1 of 1

Erratum: PrimerPooler: automated primer pooling to prepare library for targeted sequencing.

Biol Methods Protoc 2019 5;4(1):bpz015. Epub 2019 Nov 5.

[This corrects the article DOI: 10.1093/biomethods/bpx006.][This corrects the article DOI: 10.1093/biomethods/bpx006.].
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http://dx.doi.org/10.1093/biomethods/bpz015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7200919PMC
November 2019

Angioimmunoblastic T-cell lymphoma contains multiple clonal T-cell populations derived from a common TET2 mutant progenitor cell.

J Pathol 2020 03 16;250(3):346-357. Epub 2020 Jan 16.

Division of Cellular and Molecular Pathology, Department of Pathology, University of Cambridge, Cambridge, UK.

Angioimmunoblastic T-cell lymphoma (AITL) is a neoplastic proliferation of T follicular helper cells with clinical and histological presentations suggesting a role of antigenic drive in its development. Genetically, it is characterized by a stepwise acquisition of somatic mutations, with early mutations involving epigenetic regulators (TET2, DNMT3A) and occurring in haematopoietic stem cells, with subsequent changes involving signaling molecules (RHOA, VAV1, PLCG1, CD28) critical for T-cell biology. To search for evidence of potential oncogenic cooperation between genetic changes and intrinsic T cell receptor (TCR) signaling, we investigated somatic mutations and T-cell receptor β (TRB) rearrangement in 119 AITL, 11 peripheral T-cell lymphomas with T follicular helper phenotype (PTCL-TFH), and 25 PTCL-NOS using Fluidigm polymerase chain reaction (PCR) and Illumina MiSeq sequencing. We confirmed frequent TET2, DNMT3A, and RHOA mutations in AITL (72%, 34%, 61%) and PTCL-TFH (73%, 36%, 45%) and showed multiple TET2 mutations (2 or 3) in 57% of the involved AITL and PTCL-TFH. Clonal TRB rearrangement was seen in 76 cases with multiple functional rearrangements (2-4) in 18 cases (24%). In selected cases, we confirmed bi-clonal T-cell populations and further demonstrated that these independent T-cell populations harboured identical TET2 mutations by using BaseScope in situ hybridization, suggesting their derivation from a common TET2 mutant progenitor cell population. Furthermore, both T-cell populations expressed CD4. Finally, in comparison with tonsillar TFH cells, both AITL and PTCL-TFH showed a significant overrepresentation of several TRB variable family members, particularly TRBV19*01. Our findings suggest the presence of parallel neoplastic evolutions from a common TET2 mutant haematopoietic progenitor pool in AITL and PTCL-TFH, albeit to be confirmed in a large series of cases. The biased TRBV usage in these lymphomas suggests that antigenic stimulation may play an important role in predilection of T cells to clonal expansion and malignant transformation. © 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
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http://dx.doi.org/10.1002/path.5376DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7064999PMC
March 2020

Distinct genetic changes reveal evolutionary history and heterogeneous molecular grade of DLBCL with MYC/BCL2 double-hit.

Leukemia 2020 05 16;34(5):1329-1341. Epub 2019 Dec 16.

Department of Pathology, University of Cambridge, Cambridge, UK.

Using a Burkitt lymphoma-like gene expression signature, we recently defined a high-risk molecular high-grade (MHG) group mainly within germinal centre B-cell like diffuse large B-cell lymphomas (GCB-DLBCL), which was enriched for MYC/BCL2 double-hit (MYC/BCL2-DH). The genetic basis underlying MHG-DLBCL and their aggressive clinical behaviour remain unknown. We investigated 697 cases of DLBCL, particularly those with MYC/BCL2-DH (n = 62) by targeted sequencing and gene expression profiling. We showed that DLBCL with MYC/BCL2-DH, and those with BCL2 translocation, harbour the characteristic mutation signatures that are associated with follicular lymphoma and its high-grade transformation. We identified frequent MYC hotspot mutations that affect the phosphorylation site (T58) and its adjacent amino acids, which are important for MYC protein degradation. These MYC mutations were seen in a subset of cases with MYC translocation, but predominantly in those of MHG. The mutations were more frequent in double-hit lymphomas with IG as the MYC translocation partner, and were associated with higher MYC protein expression and poor patient survival. DLBCL with MYC/BCL2-DH and those with BCL2 translocation alone are most likely derived from follicular lymphoma or its precursor lesion, and acquisition of MYC pathogenic mutations may augment MYC function, resulting in aggressive clinical behaviour.
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http://dx.doi.org/10.1038/s41375-019-0691-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7192846PMC
May 2020

Utility of ctDNA to support patient selection for early phase clinical trials: the TARGET study.

Nat Med 2019 05 22;25(5):738-743. Epub 2019 Apr 22.

Experimental Cancer Medicine Team, The Christie NHS Foundation Trust, Manchester, UK.

Next-generation sequencing (NGS) of circulating tumor DNA (ctDNA) supports blood-based genomic profiling but is not yet routinely implemented in the setting of a phase I trials clinic. TARGET is a molecular profiling program with the primary aim to match patients with a broad range of advanced cancers to early phase clinical trials on the basis of analysis of both somatic mutations and copy number alterations (CNA) across a 641 cancer-associated-gene panel in a single ctDNA assay. For the first 100 TARGET patients, ctDNA data showed good concordance with matched tumor and results were turned round within a clinically acceptable timeframe for Molecular Tumor Board (MTB) review. When a 2.5% variant allele frequency (VAF) threshold was applied, actionable mutations were identified in 41 of 100 patients, and 11 of these patients received a matched therapy. These data support the application of ctDNA in this early phase trial setting where broad genomic profiling of contemporaneous tumor material enhances patient stratification to novel therapies and provides a practical template for bringing routinely applied blood-based analyses to the clinic.
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http://dx.doi.org/10.1038/s41591-019-0380-zDOI Listing
May 2019

Gene-expression profiling of bortezomib added to standard chemoimmunotherapy for diffuse large B-cell lymphoma (REMoDL-B): an open-label, randomised, phase 3 trial.

Lancet Oncol 2019 05 1;20(5):649-662. Epub 2019 Apr 1.

Cancer Research UK Centre, University of Southampton, Southampton, UK. Electronic address:

Background: Biologically distinct subtypes of diffuse large B-cell lymphoma can be identified using gene-expression analysis to determine their cell of origin, corresponding to germinal centre or activated B cell. We aimed to investigate whether adding bortezomib to standard therapy could improve outcomes in patients with these subtypes.

Methods: In a randomised evaluation of molecular guided therapy for diffuse large B-cell lymphoma with bortezomib (REMoDL-B), an open-label, adaptive, randomised controlled, phase 3 superiority trial, participants were recruited from 107 cancer centres in the UK (n=94) and Switzerland (n=13). Eligible patients had previously untreated, histologically confirmed diffuse large B-cell lymphoma with sufficient diagnostic material from initial biopsies for gene-expression profiling and pathology review; were aged 18 years or older; had ECOG performance status of 2 or less; had bulky stage I or stage II-IV disease requiring full-course chemotherapy; had measurable disease; and had cardiac, lung, renal, and liver function sufficient to tolerate chemotherapy. Patients initially received one 21-day cycle of standard rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone (R-CHOP; rituximab 375 mg/m, cyclophosphamide 750 mg/m, doxorubicin 50 mg/m, and vincristine 1·4 mg/m [to a maximum of 2 mg total dose] intravenously on day 1 of the cycle, and prednisolone 100 mg orally once daily on days 1-5). During this time, we did gene-expression profiling using whole genome cDNA-mediated annealing, selection, extension, and ligation assay of tissue from routine diagnostic biopsy samples to determine the cell-of-origin subtype of each participant (germinal centre B cell, activated B cell, or unclassified). Patients were then centrally randomly assigned (1:1) via a web-based system, with block randomisation stratified by international prognostic index score and cell-of-origin subtype, to continue R-CHOP alone (R-CHOP group; control), or with bortezomib (RB-CHOP group; experimental; 1·3 mg/m intravenously or 1·6 mg/m subcutaneously) on days 1 and 8 for cycles two to six. If RNA extracted from the diagnostic tissues was of insufficient quality or quantity, participants were given R-CHOP as per the control group. The primary endpoint was 30-month progression-free survival, for the germinal centre and activated B-cell population. The primary analysis was on the modified intention-to-treat population of activated and germinal centre B-cell population. Safety was assessed in all participants who were given at least one dose of study drug. We report the progression-free survival and safety outcomes for patients in the follow-up phase after the required number of events occurred. This study was registered at ClinicalTrials.gov, number NCT01324596, and recruitment and treatment has completed for all participants, with long-term follow-up ongoing.

Findings: Between June 2, 2011, and June 10, 2015, 1128 eligible patients were registered, of whom 918 (81%) were randomly assigned to receive treatment (n=459 to R-CHOP, n=459 to RB-CHOP), comprising 244 (26·6%) with activated B-cell disease, 475 (51·7%) with germinal centre B cell disease, and 199 (21·7%) with unclassified disease. At a median follow-up of 29·7 months (95% CI 29·0-32·0), we saw no evidence for a difference in progression-free survival in the combined germinal centre and activated B-cell population between R-CHOP and RB-CHOP (30-month progression-free survival 70·1%, 95% CI 65·0-74·7 vs 74·3%, 69·3-78·7; hazard ratio 0·86, 95% CI 0·65-1·13; p=0·28). The most common grade 3 or worse adverse event was haematological toxicity, reported in 178 (39·8%) of 447 patients given R-CHOP and 187 (42·1%) of 444 given RB-CHOP. However, RB-CHOP was not associated with increased haematological toxicity and 398 [87·1%] of 459 participants assigned to receive RB-CHOP completed six cycles of treatment. Grade 3 or worse neuropathy occurred in 17 (3·8%) patients given RB-CHOP versus eight (1·8%) given R-CHOP. Serious adverse events occurred in 190 (42·5%) patients given R-CHOP, including five treatment-related deaths, and 223 (50·2%) given RB-CHOP, including four treatment-related deaths.

Interpretation: This is the first large-scale study in diffuse large B-cell lymphoma to use real-time molecular characterisation for prospective stratification, randomisation, and subsequent analysis of biologically distinct subgroups of patients. The addition of bortezomib did not improve progression-free survival.

Funding: Janssen-Cilag, Bloodwise, and Cancer Research UK.
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http://dx.doi.org/10.1016/S1470-2045(18)30935-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6494978PMC
May 2019

Variable Responses of MYC Translocation Positive Lymphoma Cell Lines To Different Combinations of Novel Agents: Impact of BCL2 Family Protein Expression.

Transl Oncol 2018 Oct 25;11(5):1147-1154. Epub 2018 Jul 25.

Division of Cellular and Molecular Pathology, Department of Pathology, University of Cambridge, Cambridge, UK; Molecular Malignancy Laboratory, Haematopathology and Oncology Diagnostic Service, Cambridge University Hospitals NHS Foundation Trust, Cambridge, UK. Electronic address:

Several newly developed drugs including JQ1 (BET inhibitor), ABT199 (BCL2 inhibitor), and bortezomib (proteasome inhibitor) may offer novel therapeutic strategies for aggressive diffuse large B-cell lymphoma (DLBCL). We tested these drugs together with doxorubicin in a series of combinations in 16 DLBCL cell lines including 4 ABC-DLBCL (OCI-Ly3, OCI-Ly10, SUDHL2, RIVA) and 12 GCB-DLBCL lines (OCI-Ly4, OCI-Ly18, BJAB, SUDHL4, SUDHL6, SUDHL10, DB, PR1, VAL, SC1, Karpas-231, Karpas-422). Among these cell lines, ABT199 and doxorubicin, and to a lesser extent JQ1 and bortezomib, showed high variations in their ED50 values. Of the six cell lines showing high ABT199 ED50 values, four (SUDHL10, OCI-Ly4, SUDHL2, and BJAB) had no or little BCL2 expression, and SUDHL6 also displayed a low BCL2 expression. There was no association between the ED50 value of doxorubicin, JQ1 and bortezomib, and TP53/MYC/BCL2 genetic abnormalities or cell of origin subtype. A synergistic effect in all or the majority of drug combinations was seen in 11 cell lines, while an antagonistic effect in a high proportion of drug combinations was observed in the remaining 5 cell lines including the 3 (SUDHL10, OCI-Ly4, and SUDHL2) with little BCL2 expression, and additionally OCI-Ly18 and RIVA. Extensive Western blot analyses revealed high MCL1 expression in SUDHL10 and OCI-Ly4 but no apparent alterations in other cell lines. The molecular mechanism underlying the antagonistic effect of drug combinations in DLBCL is heterogeneous with the altered BCL2 family protein expression (absent BCL2, but high MCL1) in some cell lines.
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http://dx.doi.org/10.1016/j.tranon.2018.07.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6077152PMC
October 2018

Mutation screening using formalin-fixed paraffin-embedded tissues: a stratified approach according to DNA quality.

Lab Invest 2018 08 16;98(8):1084-1092. Epub 2018 May 16.

Department of Pathology, University of Cambridge, Cambridge, UK.

DNA samples from formalin-fixed paraffin-embedded tissues are highly degraded with variable quality, and this imposes a big challenge for targeted sequencing due to false positives, largely caused by PCR errors and cytosine deamination. To eliminate false positives, a common practice is to validate the detected variants by Sanger sequencing or perform targeted sequencing in duplicate. Technically, PCR errors could be removed by molecular barcoding of template DNA prior to amplification as in the HaloPlexHS design. Nonetheless, it is uncertain to what extent variants detected using this approach should be further validated. Here, we addressed this question by correlating variant reproducibility with DNA quality using HaloPlexHS target enrichment and Illumina HiSeq4000, together with an in-house validated variant calling algorithm. The overall sequencing coverage, as shown by analyses of 70 genes in 266 cases of large B-cell lymphoma, was excellent (98%) in DNA samples amenable for PCR of ≥400 bp, but suboptimal (92%) and poor (80%) in those amenable for PCR of 300 bp and 200 bp respectively. By mutation analysis in duplicate in 93 cases, we demonstrated that 20 alternative allele depth (AAD) was an optimal cut-off value for separating reproducible from non-reproducible variants in DNA samples amenable for PCR of ≥300 bp, with 97% sensitivity and 100% specificity. By cross validation with a previously established targeted sequencing protocol by Fluidigm-PCR and Illumina MiSeq, the HaloPlexHS protocol was shown to be highly sensitive and specific in mutation screening. To conclude, we proposed a stratified approach for mutation screening by HaloplexHS and Illumina HiSeq4000 according to DNA quality. DNA samples with good quality (≥400 bp) are amenable for mutation analysis with a single replicate, with only variants at 15-20 AAD requiring for further validation, while those with suboptimal quality (300 bp) are better analysed in duplicate with reproducible variants at >15 AAD regarded as true genetic changes.
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http://dx.doi.org/10.1038/s41374-018-0066-zDOI Listing
August 2018

Early loss of Crebbp confers malignant stem cell properties on lymphoid progenitors.

Nat Cell Biol 2017 Sep 21;19(9):1093-1104. Epub 2017 Aug 21.

Wellcome Trust-MRC Cambridge Stem Cell Institute, Cambridge, UK.

Loss-of-function mutations of cyclic-AMP response element binding protein, binding protein (CREBBP) are prevalent in lymphoid malignancies. However, the tumour suppressor functions of CREBBP remain unclear. We demonstrate that loss of Crebbp in murine haematopoietic stem and progenitor cells (HSPCs) leads to increased development of B-cell lymphomas. This is preceded by accumulation of hyperproliferative lymphoid progenitors with a defective DNA damage response (DDR) due to a failure to acetylate p53. We identify a premalignant lymphoma stem cell population with decreased H3K27ac, which undergoes transcriptional and genetic evolution due to the altered DDR, resulting in lymphomagenesis. Importantly, when Crebbp is lost later in lymphopoiesis, cellular abnormalities are lost and tumour generation is attenuated. We also document that CREBBP mutations may occur in HSPCs from patients with CREBBP-mutated lymphoma. These data suggest that earlier loss of Crebbp is advantageous for lymphoid transformation and inform the cellular origins and subsequent evolution of lymphoid malignancies.
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http://dx.doi.org/10.1038/ncb3597DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5633079PMC
September 2017

Significant association between TNFAIP3 inactivation and biased immunoglobulin heavy chain variable region 4-34 usage in mucosa-associated lymphoid tissue lymphoma.

J Pathol 2017 09 7;243(1):3-8. Epub 2017 Aug 7.

Division of Cellular and Molecular Pathology, Department of Pathology, University of Cambridge, Cambridge, UK.

Both antigenic drive and genetic change play critical roles in the development of mucosa-associated lymphoid tissue (MALT) lymphoma, but neither alone is sufficient for malignant transformation, and lymphoma development critically depends on their cooperation. However, which of these different events concur and how they cooperate in MALT lymphomagenesis is totally unknown. To explore this, we investigated somatic mutations of 17 genes and immunoglobulin heavy chain variable region (IGHV) usage in 179 MALT lymphomas from various sites. We showed that: (1) there was a significant association between the biased usage of IGHV4-34 (binds to the carbohydrate I/i antigens) and inactivating mutation of TNFAIP3 [encoding a global negative regulator of the canonical nuclear factor-κB (NF-κB) pathway] in ocular adnexal MALT lymphoma; (2) IGHV1-69 was significantly overrepresented (54%) in MALT lymphoma of the salivary gland, but was not associated with mutation in any of the 17 genes investigated; and (3) MALT lymphoma lacked mutations that are frequently seen in other B-cell lymphomas characterized by constitutive NF-κB activities, including mutations in CD79B, CARD11, MYD88, TNFRSF11A, and TRAF3. Our findings show, for the first time, a significant association between biased usage of autoreactive IGHV and somatic mutation of NF-κB regulators in MALT lymphoma, arguing for their cooperation in sustaining chronic B-cell receptor signalling and driving oncogenesis in lymphoma development. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/path.4933DOI Listing
September 2017

PrimerPooler: automated primer pooling to prepare library for targeted sequencing.

Biol Methods Protoc 2017 Jan 12;2(1):bpx006. Epub 2017 May 12.

Department of Pathology, Division of Cellular and Molecular Pathology, University of Cambridge, CB2 0QQ, UK.

Targeted next-generation sequencing based on PCR amplification involves pooling of hundreds to thousands of primers, for preamplification and subsequent parallel single/multiplex PCR. It is often necessary to allocate the set of primers into subpools, a common issue being potential cross-hybridization. For smaller numbers of primers, pool division can be done manually using trial and error to minimize potential hybridization, but this becomes inefficient and time consuming with increasing numbers of primer pairs. We developed PrimerPooler that automates swapping of primer pairs between any user-defined number of subpools to obtain combinations with low-potential interactions. PrimerPooler performs inter-/intra-primer hybridization analysis to identify the adverse interactions, as well as simultaneous mapping of all primers onto a genome sequence in a single run without requiring a prior index of the genome. This allows detection of overlapping primer pairs and allocation of these primer pairs into separate subpools where tiling approaches are used. Using PrimerPooler, 1153 primer pairs were assigned to three preamplification pools (388, 389 and 376 primer pairs each), then 144 subpools of six- to nine-plex PCR for Fluidigm Access Array PCR, followed by Illumina MiSeq sequencing. With optimized experimental protocols, an average of 3269 reads was achieved for the targeted regions, with 95% of targets covered by at least 50 reads, the minimal depth of reads for confident variant calling. PrimerPooler provides a fast and highly efficient stratification of primer pairs for targeted enrichment, thus ensuring representative amplification of the targeted sequences. PrimerPooler is also able to analyse degenerate primers, and is thus also useful for microbiological identification and related target sequencing.
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http://dx.doi.org/10.1093/biomethods/bpx006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6994079PMC
January 2017

The prognosis of translocation positive diffuse large B-cell lymphoma depends on the second hit.

J Pathol Clin Res 2015 Jul 30;1(3):125-133. Epub 2015 Mar 30.

Division of Molecular HistopathologyDepartment of PathologyUniversity of CambridgeUK; Department of HistopathologyAddenbrooke's Hospital, Cambridge University Hospitals NHS Foundation TrustCambridgeUK.

A proportion of translocation positive diffuse large B-cell lymphomas (DLBCL) harbour a and/or translocation, known as double-hit DLBCL, and are clinically aggressive. It is unknown whether there are other genetic abnormalities that cooperate with translocation and form double-hit DLBCL, and whether there is a difference in clinical outcome between the double-hit DLBCL and those with an isolated translocation. We investigated gene mutations along with and translocations in a total of 234 cases of DLBCL, including 81 with translocation. mutations were investigated by PCR and sequencing, while and translocation was studied by interphase fluorescence in situ hybridization. The majority of translocation positive DLBCLs (60/81 = 74%) had at least one additional genetic hit. In translocation positive DLBCL treated by R-CHOP ( = 67), mutation and but not translocation had an adverse effect on patient overall survival. In comparison with DLBCL with an isolated translocation, cases with double-hits had the worst overall survival, followed by those with double-hits. In translocation negative DLBCL treated by R-CHOP ( = 101), mutation, and translocation had no impact on patient survival. The prognosis of translocation positive DLBCL critically depends on the second hit, with mutations and translocation contributing to an adverse prognosis. It is pivotal to investigate both mutations and translocations in translocation positive DLBCL, and to distinguish double-hit DLBCLs from those with an isolated translocation.
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http://dx.doi.org/10.1002/cjp2.10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4915334PMC
July 2015

Somatic Mutation Screening Using Archival Formalin-Fixed, Paraffin-Embedded Tissues by Fluidigm Multiplex PCR and Illumina Sequencing.

J Mol Diagn 2015 Sep 9;17(5):521-32. Epub 2015 Jul 9.

Division of Molecular Histopathology, Department of Pathology, University of Cambridge, Cambridge, United Kingdom. Electronic address:

High-throughput somatic mutation screening using FFPE tissues is a major challenge because of a lack of established methods and validated variant calling algorithms. We aimed to develop a targeted sequencing protocol by Fluidigm multiplex PCR and Illumina sequencing and to establish a companion variant calling algorithm. The experimental protocol and variant calling algorithm were first developed and optimized against a series of somatic mutations (147 substitutions, 12 indels ranging from 1 to 33 bp) in seven genes, previously detected by Sanger sequencing of DNA from 163 FFPE lymphoma biopsy specimens. The optimized experimental protocol and variant calling algorithm were further ascertained in two separate experiments by including the seven genes as a part of larger gene panels (22 or 13 genes) using FFPE and high-molecular-weight lymphoma DNAs, respectively. We found that most false-positive variants were due to DNA degradation, deamination, and Taq polymerase errors, but they were nonreproducible and could be efficiently eliminated by duplicate experiments. A small fraction of false-positive variants appeared in duplicate, but they were at low alternative allele frequencies and could be separated from mutations when appropriate threshold value was used. In conclusion, we established a robust practical approach for high-throughput mutation screening using archival FFPE tissues.
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http://dx.doi.org/10.1016/j.jmoldx.2015.04.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4597275PMC
September 2015

Aggressive natural killer-cell neoplasm presenting in the marrow: a report of two cases including one with gains of chromosomes 4q and 9p.

Diagn Pathol 2015 Jul 4;10:88. Epub 2015 Jul 4.

Department of Pathology, Chi-Mei Medical Centre, 901 Chung-Hwa Road, Yung-Kang District, Tainan, Taiwan.

Aggressive nature killer (NK)-cell neoplasm includes aggressive NK-cell leukemia (ANKL) and extranodal NK/T-cell lymphoma (ENKTL), nasal type. ANKL is rare and is characterized by a systemic neoplastic proliferation of NK-cells, usually with a leukemic presentation. ENKTL is a predominantly extranodal lymphoma, occurring mainly in the upper aerodigestive tract. Both are aggressive neoplasms strongly associated with Epstein-Barr virus (EBV). Here we report two patients with aggressive NK-cells neoplasms localized in the bone marrow (BM) who presented as prolonged fever, anemia, and thrombocytopenia. Both were treated initially as infectious disease. Imaging studies revealed splenomegaly without any nodular lesion or lymphadenopathy. BM examination revealed extensive involvement by EBV-positive NK-cells in both cases. Staging workup including nasal examination/biopsy was negative. Both patients passed away in a month. One case showed gains of chromosomes 4q and 9p by array comparative genomic hybridization. Both tumors were diagnostically challenging due to the unusual clinical presentation and absence of leukemic change, tumor mass or lymphadenopathy. Our cases demonstrate that lymphoma should be considered in patients with fever of unknown origin and bone marrow aspiration/biopsy should be performed as early diagnosis and novel therapeutic regimens may benefit these patients.
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http://dx.doi.org/10.1186/s13000-015-0333-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4491245PMC
July 2015

Bivalent enzyme inhibitors discovered using dynamic covalent chemistry.

Chemistry 2012 Aug 10;18(34):10562-70. Epub 2012 Jul 10.

School of Chemistry, University of Edinburgh, King's Buildings, West Mains Rd., Edinburgh, UK.

A bivalent dynamic covalent chemistry (DCC) system has been designed to selectively target members of the homodimeric glutathione-S-transferase (GST) enzyme family. The dynamic covalent libraries (DCLs) use aniline-catalysed acylhydrazone exchange between bivalent hydrazides and glutathione-conjugated aldehydes and the bis-hydrazides act as linkers to bridge between each glutathione binding site. The resultant DCLs were found to be compatible and highly responsive to templating with different GST isozymes, with the best results coming from the M and Schistosoma japonicum (Sj) class of GSTs, targets in cancer and tropical disease, respectively. The approach yielded compounds with selective, nanomolar affinity (K(i) =61 nM for mGSTM1-1) and demonstrates that DCC can be used to simultaneously interrogate binding sites on different subunits of a dimeric protein.
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http://dx.doi.org/10.1002/chem.201201507DOI Listing
August 2012

Nucleolar targeting of RelA(p65) is regulated by COMMD1-dependent ubiquitination.

Cancer Res 2010 Jan;70(1):139-49

Colon Cancer Genetics Group, University of Edinburgh Cancer Research Centre and MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, Western General Hospital, Edinburgh, United Kingdom.

Stimulation of the NF-kappaB pathway can have proapoptotic or antiapoptotic consequences, and one mechanism that determines the outcome is the nuclear distribution of RelA. Certain stress stimuli induce nucleolar accumulation of RelA thereby mediating apoptosis, whereas others induce nucleoplasmic accumulation and inhibition of apoptosis. Here we investigated the mechanisms that regulate the nuclear distribution of RelA, specifically, the role of the ubiquitin/proteasome system. We found that stress-induced nucleolar translocation of RelA is preceded by ubiquitination of the protein. We also found that chemical proteasome inhibitors induce the ubiquitination and nucleolar translocation of RelA and that this is required for the apoptotic response to these agents. We show that the RelA nucleolar localization signal (amino acids 27-30) is a critical domain for ubiquitination of the protein but that the lysine residue within this motif is not a direct target. We show that RelA binds COMMD1, the rate-limiting component of the RelA ubiquitin ligase complex, in response to stress. Furthermore, we show that overexpression of COMMD1 promotes stress-mediated nucleolar targeting of RelA, whereas knockdown of COMMD1 blocks this effect, causing RelA to remain in the nucleoplasm. These data identify a new role for COMMD1 in regulating the nuclear/nucleolar distribution of RelA and suggest that ubiquitination acts as a signal for transport of RelA to the nucleolus. These findings have relevance to the design of chemopreventative/anticancer agents that act by targeting RelA to the nucleolar compartment.
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http://dx.doi.org/10.1158/0008-5472.CAN-09-1397DOI Listing
January 2010