Publications by authors named "Adrian Dubuc"

72 Publications

Re-evaluating tumors of purported specialized prostatic stromal origin reveals molecular heterogeneity, including non-recurring gene fusions characteristic of uterine and soft tissue sarcoma subtypes.

Mod Pathol 2021 May 13. Epub 2021 May 13.

Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.

Tumors of purported specialized prostatic stromal origin comprise prostatic stromal sarcomas (PSS) and stromal tumors of uncertain malignant potential (STUMP). Prior studies have described their clinicopathologic characteristics, but the molecular features remain incompletely understood. Moreover, these neoplasms are morphologically heterogeneous and the lack of specific adjunctive markers of prostatic stromal lineage make precise definition more difficult, leading some to question whether they represent a specific tumor type. In this study, we used next-generation DNA and RNA sequencing to profile 25 primary prostatic mesenchymal neoplasms of possible specialized prostatic stromal origin, including cases originally diagnosed as PSS (11) and STUMP (14). Morphologically, the series comprised 20 cases with solid architecture (11 PSS and 9 STUMP) and 5 cases with phyllodes-like growth pattern (all STUMP). Combined DNA and RNA sequencing results demonstrated that 19/22 (86%) cases that underwent successful sequencing (either DNA or RNA) harbored pathogenic somatic variants. Except for TP53 alterations (6 cases), ATRX mutations (2 cases), and a few copy number variants (-13q, -14q, -16q and +8/8p), the findings were largely nonrecurrent. Eight gene rearrangements were found, and 4 (NAB2-STAT6, JAZF1-SUZ12, TPM3-NTRK1 and BCOR-MAML3) were useful for reclassification of the cases as specific entities. The present study shows that mesenchymal neoplasms of the prostate are morphologically and molecularly heterogeneous and include neoplasms that harbor genetic aberrations seen in specific mesenchymal tumors arising in other anatomic sites, including soft tissue and the uterus. These data suggest that tumors of purported specialized prostatic stromal origin may perhaps not represent a single diagnostic entity or specific disease group and that alternative diagnoses should be carefully considered.
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http://dx.doi.org/10.1038/s41379-021-00818-6DOI Listing
May 2021

Near haploidization is a genomic hallmark which defines a molecular subgroup of giant cell glioblastoma.

Neurooncol Adv 2020 Jan-Dec;2(1):vdaa155. Epub 2020 Nov 12.

Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts, USA.

Background: Giant cell glioblastoma (gcGBM) is a rare histologic subtype of glioblastoma characterized by numerous bizarre multinucleate giant cells and increased reticulin deposition. Compared with conventional isocitrate dehydrogenase (IDH)-wildtype glioblastomas, gcGBMs typically occur in younger patients and are generally associated with an improved prognosis. Although prior studies of gcGBMs have shown enrichment of genetic events, such as alterations, no defining aberrations have been identified. The aim of this study was to evaluate the genomic profile of gcGBMs to facilitate more accurate diagnosis and prognostication for this entity.

Methods: Through a multi-institutional collaborative effort, we characterized 10 gcGBMs by chromosome studies, single nucleotide polymorphism microarray analysis, and targeted next-generation sequencing. These tumors were subsequently compared to the genomic and epigenomic profile of glioblastomas described in The Cancer Genome Atlas (TCGA) dataset.

Results: Our analysis identified a specific pattern of genome-wide massive loss of heterozygosity (LOH) driven by near haploidization in a subset of glioblastomas with giant cell histology. We compared the genomic signature of these tumors against that of all glioblastomas in the TCGA dataset ( = 367) and confirmed that our cohort of gcGBMs demonstrated a significantly different genomic profile. Integrated genomic and histologic review of the TCGA cohort identified 3 additional gcGBMs with a near haploid genomic profile.

Conclusions: Massive LOH driven by haploidization represents a defining molecular hallmark of a subtype of gcGBM. This unusual mechanism of tumorigenesis provides a diagnostic genomic hallmark to evaluate in future cases, may explain reported differences in survival, and suggests new therapeutic vulnerabilities.
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http://dx.doi.org/10.1093/noajnl/vdaa155DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7764500PMC
November 2020

Correlation of methylthioadenosine phosphorylase (MTAP) protein expression with MTAP and CDKN2A copy number in malignant pleural mesothelioma.

Histopathology 2021 Jun 14;78(7):1032-1042. Epub 2021 Apr 14.

Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA.

Aims: Methylthioadenosine phosphorylase (MTAP) immunohistochemical expression is a specific marker of CDKN2A deletion in malignant mesothelioma. However, the relationship of MTAP expression with MTAP copy number remains unexplored.

Methods And Results: Forty malignant pleural mesotheliomas were characterised by targeted next-generation sequencing (29), single-nucleotide polymorphism microarray (seven), or both (four). MTAP and CDKN2A copy numbers were correlated with MTAP expression. Twenty-seven (68%) tumours showed CDKN2A deletion (14 heterozygous; 13 homozygous), of which 20 (74%) showed MTAP codeletion (15 heterozygous; five homozygous). No tumours showed MTAP deletion without CDKN2A codeletion. Loss of MTAP expression was seen in 16 (40%) tumours, and was 75% sensitive and 95% specific for MTAP deletion, and 59% sensitive and 100% specific for CDKN2A deletion. Nine of 40 (23%) tumours showed heterogeneous MTAP staining, and the percentage of tumour cells with MTAP loss correlated with molecular detection of MTAP deletion.

Conclusions: MTAP is frequently codeleted with CDKN2A in pleural mesothelioma. However, homozygous deletion of both genes occurs in a minority of tumours (5/40; 13%); CDKN2A deletion often co-occurs with heterozygous MTAP deletion or neutral MTAP copy number; and MTAP expression correlates inconsistently with heterozygous MTAP deletion. Correspondingly, MTAP immunohistochemistry is a highly specific but only moderately sensitive assay for CDKN2A deletion.
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http://dx.doi.org/10.1111/his.14324DOI Listing
June 2021

Quantification of aneuploidy in targeted sequencing data using ASCETS.

Bioinformatics 2020 Nov 28. Epub 2020 Nov 28.

Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA.

Summary: The expansion of targeted panel sequencing efforts has created opportunities for large-scale genomic analysis, but tools for copy-number quantification on panel data are lacking. We introduce ASCETS, a method for the efficient quantitation of arm and chromosome-level copy-number changes from targeted sequencing data.

Availability: ASCETS is implemented in R and is freely available to non-commercial users on GitHub: https://github.com/beroukhim-lab/ascets, along with detailed documentation.

Supplementary Information: Supplementary data are available at Bioinformatics online.
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http://dx.doi.org/10.1093/bioinformatics/btaa980DOI Listing
November 2020

A Role for Chromosomal Microarray Testing in the Workup of Male Infertility.

J Mol Diagn 2020 09 29;22(9):1189-1198. Epub 2020 Jun 29.

Department of Pathology, Harvard Medical School and Brigham and Women's Hospital, Boston, Massachusetts; Laboratory for Molecular Medicine, Partners HealthCare Personalized Medicine, Cambridge, Massachusetts. Electronic address:

Genetic analysis is a critical component in the male infertility workup. For male infertility due to oligospermia/azoospermia, standard guidelines recommend karyotype and Y-chromosome microdeletion analyses. A karyotype is used to identify structural and numerical chromosome abnormalities, whereas Y-chromosome microdeletions are commonly evaluated by multiplex PCR analysis because of their submicroscopic size. Because these assays often require different Vacutainer tubes to be sent to different laboratories, ordering is prone to errors. In addition, this workflow limits the ability for sequential testing and a comprehensive test result. A potential solution includes performing Y-microdeletion and numerical chromosome analysis-the most common genetic causes of oligospermia/azoospermia-by chromosomal microarray (CMA) and reflexing to karyotype as both assays are often offered in the cytogenetics laboratory. Such analyses can be performed using one sodium heparin Vacutainer tube sample. To determine the effectiveness of CMA for the detection of clinically significant Y-chromosome microdeletions, 21 cases with known Y microdeletions were tested by CytoScan HD platform. CMA studies identified all known Y-chromosome microdeletions, and in 11 cases (52%) identified additional clinically important cytogenetic anomalies, including six cases of 46, XX males, one case of isodicentric Y, two cases of a dicentric Y, and three cases of terminal Yq deletions. These findings demonstrate that this testing strategy would simplify ordering and allow for an integrated interpretation of test results.
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http://dx.doi.org/10.1016/j.jmoldx.2020.06.009DOI Listing
September 2020

Dysplastic lipoma: potential diagnostic pitfall of using MDM2 RNA in situ hybridization to distinguish between lipoma and atypical lipomatous tumor.

Hum Pathol 2020 07 18;101:53-57. Epub 2020 May 18.

Department of Pathology, Massachusetts General Hospital and Harvard Medical School, 55 Fruit Street, Boston, MA, 02114, United States. Electronic address:

The distinction between lipoma and atypical lipomatous tumor can be challenging in some cases. While detection of MDM2 gene amplification via fluorescence in situ hybridization (FISH) has been well established as a diagnostic tool to distinguish atypical lipomatous tumor and well-differentiated liposarcoma from benign mimics, MDM2 RNA in situ hybridization (RNA-ISH) has recently been proposed as an alternative diagnostic assay. During clinical workup for lipomatous tumors using MDM2 RNA-ISH, we noticed several dysplastic lipomas that were positive for MDM2 RNA-ISH but negative for MDM2 amplification by FISH. In this study, we examined a series of 11 dysplastic lipomas, all confirmed to be negative for MDM2 amplification by FISH. Positive MDM2 RNA-ISH was noted in 10 (91%) dysplastic lipomas. Single-nucleotide polymorphism array on one dysplastic lipoma identified the presence of homozygous deletion of 13q, including the RB1 gene locus with no evidence of MDM2 copy number gain. Our findings on the discordance between MDM2 FISH and MDM2 RNA-ISH highlight the potential utility and pitfalls of using MDM2 RNA-ISH in the distinction of atypical lipomatous tumor and related liposarcomas from dysplastic lipoma.
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http://dx.doi.org/10.1016/j.humpath.2020.05.004DOI Listing
July 2020

From Banding to BAM Files: Genomics Informs Diagnosis and Precision Medicine for Brain Tumors.

Authors:
Adrian M Dubuc

Surg Pathol Clin 2020 Jun;13(2):343-347

Department of Pathology, Harvard Medical School, Brigham and Women's Hospital, 75 Francis Street, Boston, MA 02115, USA. Electronic address:

Tumors of the central nervous system (CNS) have been historically classified according to their morphologic and immunohistochemical features. In 2016, updates to the classification of tumors of the CNS by the World Health Organization revolutionized this paradigm. For the first time, genomic findings, whether whole-arm chromosomal aberrations or single nucleotide variants, represent a necessary and critical component of diagnosis, contributing or superseding histologic findings. These updates stem from decades of technical innovation and genomic discovery. During this time, there has been a dramatic expansion and evolution in clinical genomic assays for these tumors, informing diagnosis and guiding therapeutic management.
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http://dx.doi.org/10.1016/j.path.2020.02.007DOI Listing
June 2020

Modeling germline mutations in pineoblastoma uncovers lysosome disruption-based therapy.

Nat Commun 2020 04 14;11(1):1825. Epub 2020 Apr 14.

Toronto General Research Institute, University Health Network, 67 College Street, Toronto, ON, M5G 2M1, Canada.

Pineoblastoma is a rare pediatric cancer induced by germline mutations in the tumor suppressors RB1 or DICER1. Presence of leptomeningeal metastases is indicative of poor prognosis. Here we report that inactivation of Rb plus p53 via a WAP-Cre transgene, commonly used to target the mammary gland during pregnancy, induces metastatic pineoblastoma resembling the human disease with 100% penetrance. A stabilizing mutation rather than deletion of p53 accelerates metastatic dissemination. Deletion of Dicer1 plus p53 via WAP-Cre also predisposes to pineoblastoma, albeit with lower penetrance. In silico analysis predicts tricyclic antidepressants such as nortriptyline as potential therapeutics for both pineoblastoma models. Nortriptyline disrupts the lysosome, leading to accumulation of non-functional autophagosome, cathepsin B release and pineoblastoma cell death. Nortriptyline further synergizes with the antineoplastic drug gemcitabine to effectively suppress pineoblastoma in our preclinical models, offering new modality for this lethal childhood malignancy.
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http://dx.doi.org/10.1038/s41467-020-15585-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7156401PMC
April 2020

Clinical response to larotrectinib in adult Philadelphia chromosome-like ALL with cryptic ETV6-NTRK3 rearrangement.

Blood Adv 2020 01;4(1):106-111

Harvard Medical School, Boston, MA.

Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL) is a subtype of Ph-negative ALL that molecularly resembles Ph-positive ALL. It shares the adverse prognosis of Ph-positive ALL, but lacks the BCR-ABL1 fusion oncogene. Instead, Ph-like ALL is associated with alternative mutations in signaling pathways. We describe a case of Ph-like ALL that harbored 2 genomic alterations, which activated signaling, an NRASGly12Asp mutation, and an ETV6-NTRK3 rearrangement. Initially, the NRAS mutation was detected at high frequency, whereas the gene fusion was only detectable with a targeted next-generation sequencing-based fusion assay, but not by fluorescence in situ hybridization analysis. The disease failed to respond to multiagent chemotherapy but investigational CD19-directed chimeric antigen receptor T-cell therapy resulted in a complete remission. However, the leukemia relapsed after 6 weeks. Intriguingly, the NRAS mutation was extinguished during the chimeric antigen receptor T-cell therapy and did not contribute to the relapse, which was instead associated with a rise in ETV6-NTRK3. The relapsed leukemia progressed with further chemo- and immunotherapy but was controlled for 6 weeks with substantial leukemic cytoreduction using the TRK inhibitor larotrectinib. Unfortunately, recovery of normal hematopoiesis was only marginal and the patient eventually succumbed to infections. These results demonstrate that larotrectinib has clinical activity in ETV6-NTRK3-associated Ph-like ALL.
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http://dx.doi.org/10.1182/bloodadvances.2019000769DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6960464PMC
January 2020

A Novel Fusion in Pediatric Medullary Thyroid Carcinoma.

Thyroid 2019 11;29(11):1704-1707

Dana-Farber/Boston Children's Cancer and Blood Disorders Center, Harvard Medical School, Boston, Massachusetts.

Medullary thyroid carcinoma (MTC) is most commonly associated with gene mutations. fusions have rarely been described, although not previously in pediatrics and not previously partnered with in MTC or any other cancer. A 10-year-old boy with progressive stridor was found to have metastatic MTC, including lung, lymph node, and adrenal metastases. Baseline calcitonin was 6703 pg/mL. While molecular testing was pending, he was treated empirically with the investigational selective RET inhibitor, LOXO-292, without improvement. Molecular testing revealed a novel - fusion. His therapy was changed to crizotinib and then to alectinib for improved tolerability. Calcitonin decreased to 663 pg/mL after 6 days of ALK inhibition. He remains on alectinib with ongoing response. A novel - fusion has now been implicated in a pediatric case of metastatic MTC. This fusion has profound clinical sensitivity to ALK inhibitors. This report expands the spectrum of fusions seen in MTC, including the first pediatric case of translocated MTC. This novel fusion with has not previously been reported in other human cancers. Given the dramatic response to ALK inhibition in this case, identifying patients with fusion MTC has important therapeutic implications.
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http://dx.doi.org/10.1089/thy.2019.0041DOI Listing
November 2019

Characterization of molecular signatures of supratentorial ependymomas.

Mod Pathol 2020 01 2;33(1):47-56. Epub 2019 Aug 2.

Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA.

Ependymomas show poor correlation between World Health Organization grade and clinical outcome. A subgroup of supratentorial ependymomas are characterized by C11orf95-RELA fusions, presumed to be secondary to chromothripsis of chromosome 11, resulting in constitutive activation of the NF-κB signaling pathway and overexpression of cyclin D1, p65, and L1 cell adhesion molecule (L1CAM). These RELA-fused ependymomas are recognized as a separate, molecularly defined World Health Organization entity and might be associated with poor clinical outcome. In this study, we show that immunohistochemistry for NF-κB signaling components, such as L1CAM, p65, and cyclin D1, can help distinguish RELA-fused from non-RELA-fused supratentorial ependymomas. Furthermore, these three markers can reliably differentiate RELA-fused ependymomas from a variety of histologic mimics. Lastly, we report that RELA-fused ependymomas may be associated with different chromosomal copy number changes and molecular alterations compared to their non-RELA-fused counterparts, providing additional insight into the genetic pathogenesis of these tumors and potential targets for directed therapies.
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http://dx.doi.org/10.1038/s41379-019-0329-2DOI Listing
January 2020

Molecular characterization of localized pleural mesothelioma.

Mod Pathol 2020 02 1;33(2):271-280. Epub 2019 Aug 1.

Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, 02115, USA.

Localized pleural mesothelioma is a rare solitary circumscribed pleural tumor that is microscopically similar to diffuse malignant pleural mesothelioma. However, the molecular characteristics and nosologic relationship with its diffuse counterpart remain unknown. In a consecutive cohort of 1110 patients with pleural mesotheliomas diagnosed in 2005-2018, we identified six (0.5%) patients diagnosed with localized pleural mesotheliomas. We gathered clinical history, evaluated the histopathology, and in select cases performed karyotypic analysis and targeted next-generation sequencing. The cohort included three women and three men (median age 63; range 28-76), often presenting incidentally during radiologic evaluation for unrelated conditions. Neoadjuvant chemotherapy was administered in two patients. All tumors (median size 5.0 cm; range 2.7-13.5 cm) demonstrated gross circumscription (with microscopic invasion into lung, soft tissue, and/or rib in four cases), mesothelioma histology (four biphasic and two epithelioid types), and mesothelial immunophenotype. Of four patients with at least 6-month follow-up, three were alive (up to 8.9 years). Genomic characterization identified several subgroups: (1) BAP1 mutations with deletions of CDKN2A and NF2 in two tumors; (2) TRAF7 mutations in two tumors, including one harboring trisomies of chromosomes 3, 5, 7, and X; and (3) genomic near-haploidization, characterized by extensive loss of heterozygosity sparing chromosomes 5 and 7. Localized pleural mesotheliomas appear genetically heterogeneous and include BAP1-mutated, TRAF7-mutated, and near-haploid subgroups. While the BAP1-mutated subgroup is similar to diffuse malignant pleural mesotheliomas, the TRAF7-mutated subgroup overlaps genetically with adenomatoid tumors and well-differentiated papillary mesotheliomas, in which recurrent TRAF7 mutations have been described. Genomic near-haploidization, identified recently in a subset of diffuse malignant pleural mesotheliomas, suggests a novel mechanism in the pathogenesis of both localized pleural mesothelioma and diffuse malignant pleural mesothelioma. Our findings describe distinctive genetic features of localized pleural mesothelioma, with both similarities to and differences from diffuse malignant pleural mesothelioma.
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http://dx.doi.org/10.1038/s41379-019-0330-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7359734PMC
February 2020

Clinical Importance of CDKN2A Loss and Monosomy 10 in Pilocytic Astrocytoma.

Cureus 2019 May 23;11(5):e4726. Epub 2019 May 23.

Neuro-Oncology, Dana-Farber / Brigham and Women's Cancer Center, Harvard Medical School, Boston, USA.

This case of a radiation-naive patient with pilocytic astrocytoma highlights how deletions of CDKN2A (cyclin-dependent kinase Inhibitor 2A) and PTEN (phosphatase and tensin homolog) portended a poor clinical outcome. Pilocytic astrocytomas are grade 1 tumors usually occurring in children and young adults with KIAA1549-BRAF fusion defining the majority of pilocytic astrocytomas. The presence of CDKN2A and PTEN loss may be associated with aggressive biology in pilocytic astrocytoma and further studies should include comprehensive genomics in a larger series of adult pilocytic astrocytoma to evaluate this previously unreported finding. Providers need to be aware of this possibility given the potential for poor outcomes.
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http://dx.doi.org/10.7759/cureus.4726DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6649926PMC
May 2019

Technical laboratory standards for interpretation and reporting of acquired copy-number abnormalities and copy-neutral loss of heterozygosity in neoplastic disorders: a joint consensus recommendation from the American College of Medical Genetics and Genomics (ACMG) and the Cancer Genomics Consortium (CGC).

Genet Med 2019 09 29;21(9):1903-1916. Epub 2019 May 29.

Department of Pathology and Laboratory Medicine, Children's Hospital Los Angeles, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA.

The detection of acquired copy-number abnormalities (CNAs) and copy-neutral loss of heterozygosity (CN-LOH) in neoplastic disorders by chromosomal microarray analysis (CMA) has significantly increased over the past few years with respect to both the number of laboratories utilizing this technology and the broader number of tumor types being assayed. This highlights the importance of standardizing the interpretation and reporting of acquired variants among laboratories. To address this need, a clinical laboratory-focused workgroup was established to draft recommendations for the interpretation and reporting of acquired CNAs and CN-LOH in neoplastic disorders. This project is a collaboration between the American College of Medical Genetics and Genomics (ACMG) and the Cancer Genomics Consortium (CGC). The recommendations put forth by the workgroup are based on literature review, empirical data, and expert consensus of the workgroup members. A four-tier evidence-based categorization system for acquired CNAs and CN-LOH was developed, which is based on the level of available evidence regarding their diagnostic, prognostic, and therapeutic relevance: tier 1, variants with strong clinical significance; tier 2, variants with some clinical significance; tier 3, clonal variants with no documented neoplastic disease association; and tier 4, benign or likely benign variants. These recommendations also provide a list of standardized definitions of terms used in the reporting of CMA findings, as well as a framework for the clinical reporting of acquired CNAs and CN-LOH, and recommendations for how to deal with suspected clinically significant germline variants.
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http://dx.doi.org/10.1038/s41436-019-0545-7DOI Listing
September 2019

Differential Expression of PLAG1 in Apocrine and Eccrine Cutaneous Mixed Tumors: Evidence for Distinct Molecular Pathogenesis.

Am J Dermatopathol 2020 Apr;42(4):251-257

Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA.

Cutaneous mixed tumors, also known as chondroid syringomas, are benign adnexal neoplasms that share histomorphologic features with pleomorphic adenomas of the salivary gland. Recent work suggests that the similarity between these 2 tumor types extends to the molecular level because both harbor identical chromosomal rearrangements involving the PLAG1 gene. The resulting nuclear PLAG1 overexpression can be detected by immunohistochemistry and has become a useful diagnostic adjunct for both tumor types. In the skin, however, there are 2 morphologically distinct types of mixed tumor, which have been referred to as apocrine-type cutaneous mixed tumor (AMT) and eccrine-type cutaneous mixed tumor (EMT). Previous studies of PLAG1 expression in cutaneous mixed tumor did not distinguish between these types. Here, we evaluated PLAG1 expression by immunohistochemistry in a cohort of 25 cutaneous mixed tumors stratified by type. PLAG1 was overexpressed in the majority of AMT cases (14 of 16) but in none of the EMT cases (0 of 9). A second gene, HMGA2, known to be upregulated in a subset of salivary gland pleomorphic adenomas, was overexpressed in only 1 case of AMT (1 of 16) and in none of the cases of EMT (0 of 9). Our results indicate that apocrine- and eccrine-type mixed tumors are associated with different pathways of molecular pathogenesis and suggest that the emerging relationship between skin and salivary gland mixed tumors is likely limited to those of apocrine type.
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http://dx.doi.org/10.1097/DAD.0000000000001393DOI Listing
April 2020

Recurrent EP300-BCOR Fusions in Pediatric Gliomas With Distinct Clinicopathologic Features.

J Neuropathol Exp Neurol 2019 04;78(4):305-314

Department of Pathology, Boston Children's Hospital and Harvard Medical School, Boston, Massachusetts.

BCOR is an epigenetic regulator and is genetically altered by mutation, deletion, or gene fusion in a range of cancers. "Central nervous system high-grade neuroepithelial tumor with BCOR alteration" is a recently described entity with characteristic internal tandem duplications within exon 15 of the BCOR gene (hereafter: CNS HGNET-BCOR ex15 ITD). In this case series of 3 patients, we report the clinicopathologic, molecular, and methylome features of gliomas with novel EP300-BCOR in-frame gene fusions, thus expanding the spectrum of BCOR alterations seen in CNS tumors. The gliomas in this series arise in children (ages 10-18), involve the supratentorial compartment, and have an infiltrative pattern of growth and a myxoid/microcystic background with frequent psammomatous calcifications and prominent chicken-wire vessels. All 3 cases had areas with low-grade morphology and 2 of them demonstrated histologic high-grade transformation. In contrast to CNS HGNET-BCOR ex15 ITD, they lack perivascular pseudorosettes. On a t-Distributed Stochastic Neighbor Embedding plot they cluster perfectly together, away from CNS HGNET-BCOR ex15ITD, consistent with a different entity. Gliomas with EP300-BCOR fusions and high-grade histology can demonstrate relatively rapid regrowth after debulking or subtotal resection.
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http://dx.doi.org/10.1093/jnen/nlz011DOI Listing
April 2019

Insertional Mutagenesis Reveals Important Genetic Drivers of Central Nervous System Embryonal Tumors.

Cancer Res 2019 03 23;79(5):905-917. Epub 2019 Jan 23.

Division of Hematology, The Hospital for Sick Children, Toronto, Ontario, Canada.

Medulloblastoma and central nervous system primitive neuroectodermal tumors (CNS-PNET) are aggressive, poorly differentiated brain tumors with limited effective therapies. Using () transposon mutagenesis, we identified novel genetic drivers of medulloblastoma and CNS-PNET. Cross-species gene expression analyses classified -driven tumors into distinct medulloblastoma and CNS-PNET subgroups, indicating they resemble human Sonic hedgehog and group 3 and 4 medulloblastoma and CNS neuroblastoma with activation. This represents the first genetically induced mouse model of CNS-PNET and a rare model of group 3 and 4 medulloblastoma. We identified several putative proto-oncogenes including , and . Genetic manipulation of these genes demonstrated a robust impact on tumorigenesis and . We also determined that FOXR2 interacts with N-MYC, increases C-MYC protein stability, and activates FAK/SRC signaling. Altogether, our study identified several promising therapeutic targets in medulloblastoma and CNS-PNET. SIGNIFICANCE: A transposon-induced mouse model identifies several novel genetic drivers and potential therapeutic targets in medulloblastoma and CNS-PNET.
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http://dx.doi.org/10.1158/0008-5472.CAN-18-1261DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6397665PMC
March 2019

ZMYM2-FGFR1 fusion as secondary change in acute myeloid leukemia.

Leuk Lymphoma 2019 02 30;60(2):556-558. Epub 2018 Aug 30.

a Department of Pathology, Brigham and Women's Hospital , Harvard Medical School , Boston ( MA) , USA.

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http://dx.doi.org/10.1080/10428194.2018.1493733DOI Listing
February 2019

Yield of additional genetic testing after chromosomal microarray for diagnosis of neurodevelopmental disability and congenital anomalies: a clinical practice resource of the American College of Medical Genetics and Genomics (ACMG).

Genet Med 2018 10 18;20(10):1105-1113. Epub 2018 Jun 18.

Division of Genetics and Genomics, Boston Children's Hospital and Harvard Medical School, Boston, Massachusetts, USA.

Purpose: Chromosomal microarray (CMA) is recommended as the first-tier test in evaluation of individuals with neurodevelopmental disability and congenital anomalies. CMA may not detect balanced cytogenomic abnormalities or uniparental disomy (UPD), and deletion/duplications and regions of homozygosity may require additional testing to clarify the mechanism and inform accurate counseling. We conducted an evidence review to synthesize data regarding the benefit of additional testing after CMA to inform a genetic diagnosis.

Methods: The review was guided by key questions related to the detection of genomic events that may require additional testing. A PubMed search for original research articles, systematic reviews, and meta-analyses was evaluated from articles published between 1 January 1983 and 31 March 2017. Based on the key questions, articles were retrieved and data extracted in parallel with comparison of results and discussion to resolve discrepancies. Variables assessed included study design and outcomes.

Results: A narrative synthesis was created for each question to describe the occurrence of, and clinical significance of, additional diagnostic findings from subsequent testing performed after CMA.

Conclusion: These findings may be used to assist the laboratory and clinician when making recommendations about additional testing after CMA, as it impacts clinical care, counseling, and diagnosis.
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http://dx.doi.org/10.1038/s41436-018-0040-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6410698PMC
October 2018

PHOX2B is a reliable immunomarker in distinguishing peripheral neuroblastic tumours from CNS embryonal tumours.

Histopathology 2018 Sep 4;73(3):483-491. Epub 2018 Jul 4.

Department of Pathology, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA.

Aims: The PHOX2B gene regulates neuronal maturation in the brain stem nuclei associated with cardiorespiratory function and in the autonomic sympathetic and enteric nervous system. PHOX2B expression is a reliable immunomarker for peripheral neuroblastic tumours; however, no systematic evaluation of central nervous system (CNS) embryonal tumours was included in the studies. We encountered two cases in which the differential diagnosis included neuroblastoma and CNS embryonal tumour, and we hypothesised that PHOX2B immunostain would be helpful in establishing the diagnosis.

Methods And Results: PHOX2B immunostain was performed on 29 paediatric cases, with adequate controls: one retroperitoneal embryonal tumour in a child with retinoblastoma (index 1), one posterior fossa embryonal tumour in a child with a neuroblastoma (index 2), seven medulloblastomas, four atypical teratoid/rhabdoid tumours (ATRT), four retinoblastomas, six pineoblastomas, four embryonal tumours with multilayered rosettes (ETMR) and two CNS embryonal tumours, not elsewhere classified. Cell lineage immunomarkers (GFAP, OLIG2, synaptophysin, NeuN, CRX, PGP 9.5), immunosurrogates for molecular alterations (beta-catenin, INI1, Lin-28), array CGH and OncoPanel were performed as needed. Medulloblastomas, ATRTs, ETMRs, retinoblastomas and CNS embryonal tumours not elsewhere classified were essentially negative for PHOX2B. Two of six pineoblastomas had significant PHOX2B expression, while the rest were negative. Index 1 was negative for PHOX2B and PGP 9.5 and positive for CRX, consistent with retinoblastoma. Index 2 had diffuse PHOX2B expression, MYCN amplification and no copy number changes of medulloblastoma, in keeping with neuroblastoma.

Conclusion: PHOX2B antibody is helpful in distinguishing between peripheral neuroblastic and CNS embryonal tumours, which are immunonegative, with the caveat that a subset of pineoblastomas has significant expression.
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http://dx.doi.org/10.1111/his.13648DOI Listing
September 2018

Convergence of BMI1 and CHD7 on ERK Signaling in Medulloblastoma.

Cell Rep 2017 Dec;21(10):2772-2784

Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, 4 Newark Street, London E1 2AT, UK. Electronic address:

We describe molecular convergence between BMI1 and CHD7 in the initiation of medulloblastoma. Identified in a functional genomic screen in mouse models, a BMI1;CHD7 expression signature within medulloblastoma characterizes patients with poor overall survival. We show that BMI1-mediated repression of the ERK1/2 pathway leads to increased proliferation and tumor burden in primary human MB cells and in a xenograft model, respectively. We provide evidence that repression of the ERK inhibitor DUSP4 by BMI1 is dependent on a more accessible chromatin configuration in G4 MB cells with low CHD7 expression. These findings extend current knowledge of the role of BMI1 and CHD7 in medulloblastoma pathogenesis, and they raise the possibility that pharmacological targeting of BMI1 or ERK may be particularly indicated in a subgroup of MB with low expression levels of CHD7.
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http://dx.doi.org/10.1016/j.celrep.2017.11.021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5732319PMC
December 2017

Nuclear CRX and FOXJ1 Expression Differentiates Non-Germ Cell Pineal Region Tumors and Supports the Ependymal Differentiation of Papillary Tumor of the Pineal Region.

Am J Surg Pathol 2017 Oct;41(10):1410-1421

*Department of Pathology, Brigham and Women's Hospital, Harvard Medical School ‡Department of Pathology, Boston Children's Hospital, Harvard Medical School Departments of §Medical Oncology ¶Cancer Biology, Dana Farber Cancer Institute, Harvard Medical School, Boston, MA †Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO ∥East Pathology and Neuropathology Center, East Hospital Group, University Hospital of Lyon Bron, France.

Papillary tumor of the pineal region (PTPR) is a neuroepithelial neoplasm first described in 2003. Despite the anatomic association of PTPR with the pineal gland, the features of these tumors resemble those of the ependymal circumventricular subcommissural organ (SCO) of the posterior third ventricle. Given the presumed distinct derivation of PTPR and pineal parenchymal tumors, we hypothesized that expression of lineage-specific transcription factors could distinguish these tumors and provide additional insight into the differentiation of PTPR. A broad series of pineal region samples was reviewed, including 7 benign pineal glands, 4 pineal cysts, 13 pineocytomas, 28 pineal parenchymal tumors of intermediate differentiation, 11 pineoblastomas, and 18 PTPR. All samples were evaluated by immunohistochemistry for expression of CRX, a master transcriptional regulator of photoreceptor differentiation expressed in pineal gland and retina and/or FOXJ1, a master transcriptional regulator of ciliogenesis expressed in normal ependymal cells and ependymal neoplasms. Diffuse nuclear CRX expression is present in 100% of pineal samples. FOXJ1 is negative in all pineal samples. CRX staining is present in 53% of PTPR, though expression is nearly always limited to rare cells. Diffuse nuclear FOXJ1 expression is present in 100% of PTPR. Fetal human SCO diffusely expressed FOXJ1 but was negative for CRX. Immunohistochemistry for FOXJ1 and CRX differentiates non-germ cell pineal region tumors with high sensitivity and specificity, including pineal parenchymal tumors and PTPR. Our findings support the hypothesis that PTPR have ependymal differentiation and are phenotypically more similar to SCO than pineal gland.
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http://dx.doi.org/10.1097/PAS.0000000000000903DOI Listing
October 2017

The current state of clinical interpretation of sequence variants.

Curr Opin Genet Dev 2017 Feb 31;42:33-39. Epub 2017 Jan 31.

Laboratory for Molecular Medicine, Partners HealthCare Personalized Medicine, 65 Landsdowne Str., Cambridge, MA 02115 USA; Department of Pathology, Harvard Medical School and Brigham and Women's Hospital, 75 Francis Str., Boston, MA 02115 USA. Electronic address:

Accurate and consistent variant classification is required for Precision Medicine. But clinical variant classification remains in its infancy. While recent guidelines put forth jointly by the American College of Medical Genetics and Genomics (ACMG) and Association of Molecular Pathology (AMP) for the classification of Mendelian variants has advanced the field, the degree of subjectivity allowed by these guidelines can still lead to inconsistent classification across clinical molecular genetic laboratories. In addition, there are currently no such guidelines for somatic cancer variants, only published institutional practices. Additional variant classification guidelines, including disease- or gene-specific criteria, along with inter-laboratory data sharing is critical for accurate and consistent variant interpretation.
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http://dx.doi.org/10.1016/j.gde.2017.01.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5446800PMC
February 2017

Clinical targeted exome-based sequencing in combination with genome-wide copy number profiling: precision medicine analysis of 203 pediatric brain tumors.

Neuro Oncol 2017 Jul;19(7):986-996

Dana-Farber/Boston Children's Cancer and Blood Disorders Center, Department of Pathology, Department of Radiology, Department of Neurosurgery, Boston Children's Hospital, Boston, Massachusetts; Department of Medical Oncology, Oncologic Pathology, Department of Pediatric Oncology, Department of Cancer Biology, Center for Cancer Genome Discovery, Dana-Farber Cancer Institute, Boston, Massachusetts; Department of Pathology, Department of Neurosurgery, Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts; Pratiti Bandopadhayay, Broad Institute of MIT and Harvard, Boston, Massachusetts; Harvard Medical School, Boston, Massachusetts.

Background: Clinical genomics platforms are needed to identify targetable alterations, but implementation of these technologies and best practices in routine clinical pediatric oncology practice are not yet well established.

Methods: Profile is an institution-wide prospective clinical research initiative that uses targeted sequencing to identify targetable alterations in tumors. OncoPanel, a multiplexed targeted exome-sequencing platform that includes 300 cancer-causing genes, was used to assess single nucleotide variants and rearrangements/indels. Alterations were annotated (Tiers 1-4) based on clinical significance, with Tier 1 alterations having well-established clinical utility. OncoCopy, a clinical genome-wide array comparative genomic hybridization (aCGH) assay, was also performed to evaluate copy number alterations and better define rearrangement breakpoints.

Results: Cancer genomes of 203 pediatric brain tumors were profiled across histological subtypes, including 117 samples analyzed by OncoPanel, 146 by OncoCopy, and 60 tumors subjected to both methodologies. OncoPanel revealed clinically relevant alterations in 56% of patients (44 cancer mutations and 20 rearrangements), including BRAF alterations that directed the use of targeted inhibitors. Rearrangements in MYB-QKI, MYBL1, BRAF, and FGFR1 were also detected. Furthermore, while copy number profiles differed across histologies, the combined use of OncoPanel and OncoCopy identified subgroup-specific alterations in 89% (17/19) of medulloblastomas.

Conclusion: The combination of OncoPanel and OncoCopy multiplex genomic assays can identify critical diagnostic, prognostic, and treatment-relevant alterations and represents an effective precision medicine approach for clinical evaluation of pediatric brain tumors.
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http://dx.doi.org/10.1093/neuonc/now294DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5570190PMC
July 2017

Institutional implementation of clinical tumor profiling on an unselected cancer population.

JCI Insight 2016 11 17;1(19):e87062. Epub 2016 Nov 17.

Mirati Therapeutics, San Diego, California.

Comprehensive genomic profiling of a patient's cancer can be used to diagnose, monitor, and recommend treatment. Clinical implementation of tumor profiling in an enterprise-wide, unselected cancer patient population has yet to be reported. We deployed a hybrid-capture and massively parallel sequencing assay (OncoPanel) for all adult and pediatric patients at our combined cancer centers. Results were categorized by pathologists based on actionability. We report the results for the first 3,727 patients tested. Our cohort consists of cancer patients unrestricted by disease site or stage. Across all consented patients, half had sufficient and available (>20% tumor) material for profiling; once specimens were received in the laboratory for pathology review, 73% were scored as adequate for genomic testing. When sufficient DNA was obtained, OncoPanel yielded a result in 96% of cases. 73% of patients harbored an actionable or informative alteration; only 19% of these represented a current standard of care for therapeutic stratification. The findings recapitulate those of previous studies of common cancers but also identify alterations, including in and , associated with response to targeted therapies. In rare cancers, potentially actionable alterations suggest the utility of a "cancer-agnostic" approach in genomic profiling. Retrospective analyses uncovered contextual genomic features that may inform therapeutic response and examples where diagnoses revised by genomic profiling markedly changed clinical management. Broad sequencing-based testing deployed across an unselected cancer cohort is feasible. Genomic results may alter management in diverse scenarios; however, additional barriers must be overcome to enable precision cancer medicine on a large scale. This work was supported by DFCI, BWH, and the National Cancer Institute (5R33CA155554 and 5K23CA157631).
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http://dx.doi.org/10.1172/jci.insight.87062DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5111542PMC
November 2016

Norrin/Frizzled4 signalling in the preneoplastic niche blocks medulloblastoma initiation.

Elife 2016 11 8;5. Epub 2016 Nov 8.

Regenerative Medicine Program, Ottawa Hospital Research Institute, Ottawa, Canada.

The tumor microenvironment is a critical modulator of carcinogenesis; however, in many tumor types, the influence of the stroma during preneoplastic stages is unknown. Here we explored the relationship between pre-tumor cells and their surrounding stroma in malignant progression of the cerebellar tumor medulloblastoma (MB). We show that activation of the vascular regulatory signalling axis mediated by Norrin (an atypical Wnt)/Frizzled4 (Fzd4) inhibits MB initiation in the mouse model. Loss of Norrin/Fzd4-mediated signalling in endothelial cells, either genetically or by short-term blockade, increases the frequency of pre-tumor lesions and creates a tumor-permissive microenvironment at the earliest, preneoplastic stages of MB. This pro-tumor stroma, characterized by angiogenic remodelling, is associated with an accelerated transition from preneoplasia to malignancy. These data expose a stromal component that regulates the earliest stages of tumorigenesis in the cerebellum, and a novel role for the Norrin/Fzd4 axis as an endogenous anti-tumor signal in the preneoplastic niche.
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http://dx.doi.org/10.7554/eLife.16764DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5100999PMC
November 2016

Genomic characterization of recurrent high-grade astroblastoma.

Cancer Genet 2016 Jul-Aug;209(7-8):321-30. Epub 2016 Jun 21.

Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA; Center for Molecular Oncologic Pathology, Dana Farber Cancer Institute, Boston, MA, USA. Electronic address:

Astroblastomas are rare primary brain tumors, diagnosed based on histologic features. Not currently assigned a WHO grade, they typically display indolent behavior, with occasional variants taking a more aggressive course. We characterized the immunohistochemical characteristics, copy number (high-resolution array comparative genomic hybridization, OncoCopy) and mutational profile (targeted next-generation exome sequencing, OncoPanel) of a cohort of seven biopsies from four patients to identify recurrent genomic events that may help distinguish astroblastomas from other more common high-grade gliomas. We found that tumor histology was variable across patients and between primary and recurrent tumor samples. No common molecular features were identified among the four tumors. Mutations commonly observed in astrocytic tumors (IDH1/2, TP53, ATRX, and PTEN) or ependymoma were not identified. However one case with rapid clinical progression displayed mutations more commonly associated with GBM (NF1(N1054H/K63)*, PIK3CA(R38H) and ERG(A403T)). Conversely, another case, originally classified as glioblastoma with nine-year survival before recurrence, lacked a GBM mutational profile. Other mutations frequently seen in lower grade gliomas (BCOR, BCORL1, ERBB3, MYB, ATM) were also present in several tumors. Copy number changes were variable across tumors. Our findings indicate that astroblastomas have variable growth patterns and morphologic features, posing significant challenges to accurate classification in the absence of diagnostically specific copy number alterations and molecular features. Their histopathologic overlap with glioblastoma will likely confound the observation of long-term GBM "survivors". Further genomic profiling is needed to determine whether these tumors represent a distinct entity and to guide management strategies.
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http://dx.doi.org/10.1016/j.cancergen.2016.06.002DOI Listing
May 2017

FISHing in the dark: How the combination of FISH and conventional karyotyping improves the diagnostic yield in CpG-stimulated chronic lymphocytic leukemia.

Am J Hematol 2016 10 22;91(10):978-83. Epub 2016 Jul 22.

Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts.

Despite significant advances in molecular genetic approaches, fluorescence in situ hybridization (FISH) remains the gold standard for the diagnostic evaluation of genomic aberrations in patients with chronic lymphocytic leukemia (CLL). Efforts to improve the diagnostic utility of molecular cytogenetic testing have led to the expansion of the traditional 4-probe CLL FISH panel. Not only do these efforts increase the cost of testing, they remain hindered by the inherent limitations of FISH studies - namely the inability to evaluate genomic changes outside of the targeted loci. While array-based profiling and next generation sequencing (NGS) have critically expanded our understanding of the molecular pathogenesis of CLL, these methodologies are not routinely used by diagnostic laboratories to evaluate copy number changes or the mutational profile of this disease. Mitogenic stimulation of CLL specimens with CpG-oligonucleotide (CpG-ODN) has been identified as a reliable and reproducible means of obtaining a karyotype, facilitating a low-resolution genome-wide analysis. Across a cohort of 1255 CpG-ODN-stimulated CLL specimens, we describe the clinical utility associated with the combinatorial use of FISH and karyotyping. Our testing algorithm achieves a higher diagnostic yield (∼10%) through the detection of complex karyotypes, well-characterized chromosomal aberrations not covered by the traditional CLL FISH panel and through the detection of concurrent secondary malignancies. Moreover, the single cell nature of this approach permits the evaluation of emerging new clinical concepts including clonal dynamics and clonal evolution. This approach can be broadly applied by diagnostic laboratories to improve the utility of traditional and molecular cytogenetic studies of CLL. Am. J. Hematol. 91:978-983, 2016. © 2016 Wiley Periodicals, Inc.
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http://dx.doi.org/10.1002/ajh.24452DOI Listing
October 2016

Clonal evolution in patients with chronic lymphocytic leukaemia developing resistance to BTK inhibition.

Nat Commun 2016 05 20;7:11589. Epub 2016 May 20.

Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.

Resistance to the Bruton's tyrosine kinase (BTK) inhibitor ibrutinib has been attributed solely to mutations in BTK and related pathway molecules. Using whole-exome and deep-targeted sequencing, we dissect evolution of ibrutinib resistance in serial samples from five chronic lymphocytic leukaemia patients. In two patients, we detect BTK-C481S mutation or multiple PLCG2 mutations. The other three patients exhibit an expansion of clones harbouring del(8p) with additional driver mutations (EP300, MLL2 and EIF2A), with one patient developing trans-differentiation into CD19-negative histiocytic sarcoma. Using droplet-microfluidic technology and growth kinetic analyses, we demonstrate the presence of ibrutinib-resistant subclones and estimate subclone size before treatment initiation. Haploinsufficiency of TRAIL-R, a consequence of del(8p), results in TRAIL insensitivity, which may contribute to ibrutinib resistance. These findings demonstrate that the ibrutinib therapy favours selection and expansion of rare subclones already present before ibrutinib treatment, and provide insight into the heterogeneity of genetic changes associated with ibrutinib resistance.
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http://dx.doi.org/10.1038/ncomms11589DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4876453PMC
May 2016