Peripheral T-cell lymphomas expressing CD30 and CD15 expand the spectrum of anaplastic large cell lymphoma, ALK-negative.

Peripheral T-cell lymphomas (PTCL) are morphologically and biologically heterogeneous and a subset expresses CD30, including anaplastic large cell lymphomas (ALCL) and a minority of PTCL, not otherwise specified (PTCL, NOS). ALCL with ALK translocations (ALCL, ALK+) are readily identified by routine diagnostic methods, but differentiating ALCL without ALK translocation (ALCL, ALK-) and PTCL, NOS expressing CD30 (PTCL CD30+) can be challenging. Furthermore, rare PTCL co-express CD30 and CD15 (PTCL CD30+CD15+); some resemble ALCL, ALK- while others resemble classic Hodgkin lymphoma. To explore the relationship between PTCL CD30+CD15+ and ALCL, ALK-, we analysed 19 cases of PTCL with CD30 expression, previously diagnosed as ALCL, ALK- (nine cases) and PTCL CD30+CD15+ (10 cases) for DUSP22/IRF4 rearrangements, coding RNA expression and selected transcriptome analysis using the NanoString nCounter gene expression analysis platform. Unsupervised clustering showed no clear segregation between ALCL, ALK- and PTCL CD30+CD15+. Three cases previously classified as PTCL CD30+CD15+ showed DUSP22/IRF4 rearrangements, favouring a diagnosis of ALCL, ALK-. Our results suggest that cases previously designated PTCL CD30+CD15+, likely fall within the spectrum of ALCL, ALK-; additionally, a subset of ALCL, ALK- with DUSP22/IRF4 rearrangement expresses CD15, consistent with previous reports and expands the immunophenotypic spectrum of this lymphoma subgroup.

Follicle Center Lymphoma (FCL) of the Lower Female Genital Tract (LFGT): A Novel Variant of Primary Cutaneous Follicle Center Lymphoma (PCFCL).

Primary cutaneous follicle center lymphoma has been distinguished from nodal follicular lymphoma (FL) based on genomic and clinical features. The nature of other extranodal FLs is not well defined. We report 15 cases of follicle center lymphoma involving the lower female genital tract. Cases were evaluated using an immunohistochemical panel for B-cell lymphoma, B-cell clonality, fluorescence in situ hybridization for BCL2 gene rearrangement, and next-generation sequencing. All patients had localized disease with no evidence of bone marrow involvement. Most cases (12/15, 80%) had a follicular pattern, at least focally. Large centrocytes were a prominent feature leading to concern for diffuse large B-cell lymphoma by referring pathologists. Neoplastic cells were positive for CD20 and BCL-6, while BCL-2 was positive in 2/15 (13%) cases. Fluorescence in situ hybridization for BCL2 gene rearrangement was negative in 10/11 (91%) cases. Next-generation sequencing performed in 10 cases revealed TNFRSF14 as the most frequently mutated gene in 6/10 (60%) cases. No case had CREBBP or KMT2D mutations as seen in nodal FL. None of the patients had progressive disease with durable complete remission achieved in 10/12 (83%) cases. The median follow-up period was 7.8 years (range: 0.2 to 20.5 y) with a 5-year overall survival of 100%. We conclude that follicle center lymphoma of the lower female genital tract is a novel variant of primary cutaneous follicle center lymphoma. Despite a frequent component of large cells, it is characterized by localized disease and low risk for dissemination. Awareness and recognition are important to distinguish these lesions from aggressive B-cell lymphomas.

Clinicopathologic and Molecular Characterization of Epstein-Barr Virus-positive Plasmacytoma.

Epstein-Barr virus (EBV)-positive plasmacytoma is a rare plasma cell neoplasm. It remains unclear whether EBV-positive plasmacytoma represents a distinct entity or a variant of plasmacytoma. It shares morphologic features with plasmablastic lymphoma (PBL) and may cause diagnostic uncertainty. To better understand EBV-positive plasmacytoma and explore diagnostic criteria, this study describes 19 cases of EBV-positive plasmacytoma, compared with 27 cases of EBV-negative plasmacytoma and 48 cases of EBV-positive PBL. We reviewed the clinicopathologic findings and performed immunohistochemistry, in situ hybridization for EBV, fluorescence in situ hybridization for MYC , and next-generation sequencing. We found that 63.2% of patients with EBV-positive plasmacytoma were immunocompromised. Anaplastic features were observed in 7/19 cases. MYC rearrangement was found in 25.0% of them, and extra copies of MYC in 81.3%. EBV-positive and EBV-negative plasmacytomas possessed similar clinicopathologic features, except more frequent cytologic atypia, bone involvement and MYC aberrations in the former group. The survival rate of patients with EBV-positive plasmacytoma was comparable to that of patients with EBV-negative plasmacytoma. In comparison to PBL, EBV-positive plasmacytoma is less commonly associated with a "starry-sky" appearance, necrosis, absence of light chain expression, and a high Ki67 index (>75%). The most recurrently mutated genes/signaling pathways in EBV-positive plasmacytoma are epigenetic regulators, MAPK pathway, and DNA damage response, while the most frequently reported mutations in PBL are not observed. Collectively, EBV-positive plasmacytoma should be regarded as a biological variant of plasmacytoma. Thorough morphologic examination remains the cornerstone for distinguishing EBV-positive plasmacytoma and PBL, and molecular studies can be a valuable complementary tool.

Congenital spindle cell rhabdomyosarcoma: An international cooperative analysis.

BACKGROUND: Spindle cell rhabdomyosarcoma (RMS) is a rare variant of RMS accounting for up to 10% of cases in infants. In older children and adults, spindle cell RMS is associated with MYOD1 mutations and a poor prognosis. In infants, it is associated with recurring fusions involving NCOA2 and VGLL2. Reports in the literature suggest a favorable prognosis for this subset, however, little is known about treatment and outcome data of infants with spindle cell RMS. METHODS: Characteristics, treatment, and outcome of an international cohort of 40 patients aged ≤ 12 months with spindle cell RMS treated from 1997 to 2018 were evaluated. RESULTS: Localized disease (LD) was diagnosed in 39 patients. The median age at diagnosis was 2.5 months (range 0-12 months). Expert pathologic review confirmed the diagnosis of spindle cell RMS in all patients. Among 26 tumors that had molecular evaluation, 13 had rearrangements of NCOA and/or VGLL. Multimodal treatment of infants with LD included conventional (age adjusted) chemotherapy (n = 37), resection (n = 31) and radiotherapy (RT) (n = 5, brachytherapy in 3). Complete remission was achieved in 37/39 patients. Progressive disease occurred in two infants, relapsed disease in three. Microscopically complete surgical resection was associated with five-year event-free survival (EFS) and overall survival (OS) of 100%. Two patients with tumors ≤ 5 cm were treated with microscopically complete resection only and were alive 1 and 4.2 years after diagnosis. The 5-year EFS and OS for infants with LD were 86% (±11; CI 95%) and 91% (±9; CI 95%), respectively. One patient had metastatic disease (NCOA fusion positive) with primary tumor in head and neck and brain metastases. This patient died despite chemotherapy and delayed resection of the primary tumor due to respiratory failure secondary to cytomegalovirus infection 1.2 years after diagnosis. CONCLUSION: Infants with spindle cell RMS have an excellent prognosis. Multimodal treatment including microscopically complete resection of the tumor is strongly recommended.

Hypoxia-activated neuropeptide Y/Y5 receptor/RhoA pathway triggers chromosomal instability and bone metastasis in Ewing sarcoma.

Adverse prognosis in Ewing sarcoma (ES) is associated with the presence of metastases, particularly in bone, tumor hypoxia and chromosomal instability (CIN). Yet, a mechanistic link between these factors remains unknown. We demonstrate that in ES, tumor hypoxia selectively exacerbates bone metastasis. This process is triggered by hypoxia-induced stimulation of the neuropeptide Y (NPY)/Y5 receptor (Y5R) pathway, which leads to RhoA over-activation and cytokinesis failure. These mitotic defects result in the formation of polyploid ES cells, the progeny of which exhibit high CIN, an ability to invade and colonize bone, and a resistance to chemotherapy. Blocking Y5R in hypoxic ES tumors prevents polyploidization and bone metastasis. Our findings provide evidence for the role of the hypoxia-inducible NPY/Y5R/RhoA axis in promoting genomic changes and subsequent osseous dissemination in ES, and suggest that targeting this pathway may prevent CIN and disease progression in ES and other cancers rich in NPY and Y5R.

Germline SUCLG2 Variants in Patients With Pheochromocytoma and Paraganglioma.

BACKGROUND: Pheochromocytoma and paraganglioma (PPGL) are neuroendocrine tumors with frequent mutations in genes linked to the tricarboxylic acid cycle. However, no pathogenic variant has been found to date in succinyl-CoA ligase (SUCL), an enzyme that provides substrate for succinate dehydrogenase (SDH; mitochondrial complex II [CII]), a known tumor suppressor in PPGL. METHODS: A cohort of 352 patients with apparently sporadic PPGL underwent genetic testing using a panel of 54 genes developed at the National Institutes of Health, including the SUCLG2 subunit of SUCL. Gene deletion, succinate levels, and protein levels were assessed in tumors where possible. To confirm the possible mechanism, we used a progenitor cell line, hPheo1, derived from a human pheochromocytoma, and ablated and re-expressed SUCLG2. RESULTS: We describe 8 germline variants in the guanosine triphosphate-binding domain of SUCLG2 in 15 patients (15 of 352, 4.3%) with apparently sporadic PPGL. Analysis of SUCLG2-mutated tumors and SUCLG2-deficient hPheo1 cells revealed absence of SUCLG2 protein, decrease in the level of the SDHB subunit of SDH, and faulty assembly of the complex II, resulting in aberrant respiration and elevated succinate accumulation. CONCLUSIONS: Our study suggests SUCLG2 as a novel candidate gene in the genetic landscape of PPGL. Large-scale sequencing may uncover additional cases harboring SUCLG2 variants and provide more detailed information about their prevalence and penetrance.

Phase I Clinical Trial of an Autologous Dendritic Cell Vaccine Against HER2 Shows Safety and Preliminary Clinical Efficacy.

BACKGROUND: Despite recent advances, there is an urgent need for agents targeting HER2-expressing cancers other than breast cancer. We report a phase I study (NCT01730118) of a dendritic cell (DC) vaccine targeting HER2 in patients with metastatic cancer or bladder cancer at high risk of relapse. PATIENTS AND METHODS: Part 1 of the study enrolled patients with HER2-expressing metastatic cancer that had progressed after at least standard treatment and patients who underwent definitive treatment for invasive bladder cancer with no evidence of disease at the time of enrollment. Part 2 enrolled patients with HER2-expressing metastatic cancer who had progressed after anti-HER2 therapy. The DC vaccines were prepared from autologous monocytes and transduced with an adenoviral vector expressing the extracellular and transmembrane domains of HER2 (AdHER2). A total of five doses were planned, and adverse events were recorded in patients who received at least one dose. Objective response was evaluated by unidimensional immune-related response criteria every 8 weeks in patients who received at least two doses. Humoral and cellular immunogenicity were assessed in patients who received more than three doses. RESULTS: A total of 33 patients were enrolled at four dose levels (5 × 106, 10 × 106, 20 × 106, and 40 × 106 DCs). Median follow-up duration was 36 weeks (4-124); 10 patients completed five doses. The main reason for going off-study was disease progression. The main adverse events attributable to the vaccine were injection-site reactions. No cardiac toxicity was noted. Seven of 21 evaluable patients (33.3%) demonstrated clinical benefit (1 complete response, 1 partial response, and 5 stable disease). After ≥3 doses, an antibody response was detected in 3 of 13 patients (23.1%), including patients with complete and partial responses. Lymphocytes from 10 of 11 patients (90.9%) showed induction of anti-HER2 responses measured by the production of at least one of interferon-gamma, granzyme B, or tumor necrosis factor-alpha, and there were multifunctional responses in 8 of 11 patients (72.7%). CONCLUSIONS: The AdHER2 DC vaccine showed evidence of immunogenicity and preliminary clinical benefit in patients with HER2-expressing cancers, along with an excellent safety profile. It shows promise for further clinical applications, especially in combination regimens.

High level MYCN amplification and distinct methylation signature define an aggressive subtype of spinal cord ependymoma.

We report a novel group of clinically aggressive spinal cord ependymomas characterized by Grade III histology, MYCN amplification, an absence of NF2 alterations or other recurrent pathogenic mutations, and a unique methylation classifier profile. Seven cases were found to have MYCN amplification in the course of routine mutational profiling of 552 patients with central nervous system tumors between December 2016 and July of 2019 and an eighth patient was identified from an unrelated set of cases. Methylation array analysis revealed that none of the 8 cases clustered with any of the nine previously described ependymoma methylation subgroups, and 7 of 8 formed their own tight unique cluster. Histologically all cases showed grade III features, and all demonstrated aggressive clinical behavior. These findings are presented in the context of data from three other studies describing similar cases. Therefore, a combined total of 27 MYCN amplified spinal cord ependymoma cases have now been reported in the literature, warranting their consideration as a distinctive subtype of spinal cord ependymoma (SP-EPN-MYCN) with their unique molecular characteristics and aggressive clinical behavior.

In vivo modeling of metastatic human high-grade serous ovarian cancer in mice.

Metastasis is responsible for 90% of human cancer mortality, yet it remains a challenge to model human cancer metastasis in vivo. Here we describe mouse models of high-grade serous ovarian cancer, also known as high-grade serous carcinoma (HGSC), the most common and deadliest human ovarian cancer type. Mice genetically engineered to harbor Dicer1 and Pten inactivation and mutant p53 robustly replicate the peritoneal metastases of human HGSC with complete penetrance. Arising from the fallopian tube, tumors spread to the ovary and metastasize throughout the pelvic and peritoneal cavities, invariably inducing hemorrhagic ascites. Widespread and abundant peritoneal metastases ultimately cause mouse deaths (100%). Besides the phenotypic and histopathological similarities, mouse HGSCs also display marked chromosomal instability, impaired DNA repair, and chemosensitivity. Faithfully recapitulating the clinical metastases as well as molecular and genomic features of human HGSC, this murine model will be valuable for elucidating the mechanisms underlying the development and progression of metastatic ovarian cancer and also for evaluating potential therapies.

Melanoma With Loss of BAP1 Expression in Patients With No Family History of BAP1-Associated Cancer Susceptibility Syndrome: A Case Series.

The presence of multiple BAP1-negative melanocytic neoplasms is a hallmark of familial cancer susceptibility syndrome caused by germline mutations in BAP1. Melanocytic tumors lacking BAP1 expression may also present as sporadic lesions in patients lacking a germline BAP1 mutation. Here, we report histomorphologic and clinical characteristics of cutaneous melanomas with loss of BAP1 expression in 4 patients with no known history of BAP1-associated cancer susceptibility syndrome. The lesions were nodular melanomas composed predominantly of intradermal large epithelioid (Spitzoid) melanocytes with nuclear pseudoinclusions as well as scattered multinucleated cells, arising in association with a typical intradermal nevus. Of the 4 patients, only 1 had recurrence. This patient had multiple recurrences with in-transit and regional lymph node metastases. To the best of our knowledge, this is the first reported series of cutaneous melanomas with loss of BAP1 expression arising in patients without a family history of cancer.

A novel splicing site IRP1 somatic mutation in a patient with pheochromocytoma and JAK2V617F positive polycythemia vera: a case report.

BACKGROUND: The role of the hypoxia signaling pathway in the pathogenesis of pheochromocytoma/paraganglioma (PPGL)-polycythemia syndrome has been elucidated. Novel somatic mutations in hypoxia-inducible factor type 2A (HIF2A) and germline mutations in prolyl hydroxylase type 1 and type 2 (PHD1 and PHD2) have been identified to cause upregulation of the hypoxia signaling pathway and its target genes including erythropoietin (EPO) and its receptor (EPOR). However, in a minority of patients presenting with this syndrome, the genetics and molecular pathogenesis remain unexplained. The aim of the present study was to uncover novel genetic causes of PPGL-polycythemia syndrome. CASE PRESENTATION: A female presented with a history of JAK2V617F positive PV, diagnosed in 2007, and right adrenal pheochromocytoma diagnosed and resected in 2011. Her polycythemia symptoms and hematocrit levels continued to worsen from 2007 to 2011, with an increased frequency of phlebotomies. Postoperatively, until early 2013, her hematocrit levels remained normalized. Following this, the hematocrit levels ranged between 46.4 and 48.9% [35-45%]. Tumor tissue from the patient was further tested for mutations in genes related to upregulation of the hypoxia signaling pathway including iron regulatory protein 1 (IRP1), which is a known regulator of HIF-2α mRNA translation. Functional studies were performed to investigate the consequences of these mutations, especially their effect on the HIF signaling pathway and EPO. Indel mutations (c.267-1_267delGGinsTA) were discovered at the exon 3 splicing site of IRP1. Minigene construct and splicing site analysis showed that the mutation led to a new splicing site and a frameshift mutation of IRP1, which caused a truncated protein. Fluorescence in situ hybridization analysis demonstrated heterozygous IRP1 deletions in tumor cells. Immunohistochemistry results confirmed the truncated IRP1 and overexpressed HIF-2α, EPO and EPOR in tumor cells. CONCLUSIONS: This is the first report which provides direct molecular genetic evidence of association between a somatic IRP1 loss-of-function mutation and PHEO and secondary polycythemia. In patients diagnosed with PHEO/PGL and polycythemia with negative genetic testing for mutations in HIF2A, PHD1/2, and VHL, IRP1 should be considered as a candidate gene.

Melanoma in patients with GATA2 deficiency.

GATA2 deficiency is a recently described genetic disorder affecting hematopoietic stem cells and is associated with immunodeficiency, hematologic malignancy, and various cutaneous pathologies including cutaneous tumors. To explore the incidence and clinical course of melanoma in patients with germline GATA2 deficiencies, we conducted a retrospective chart review of 71 such patients and identified two with invasive melanoma. One melanoma was diagnosed early because it was associated with pruritus due to a graft-versus-tumor effect following bone marrow transplantation. The other one, a lentigo maligna melanoma, was locally excised but progressed to widespread metastasis and death several years later. Our observations and published studies of melanoma biology suggest an association between decreased GATA2 expression and melanoma progression. These findings suggest that GATA2 deficient patients may have an increased risk of melanoma and should be observed closely for new or changing skin lesions.

Correlation between ERG Fusion Protein and Androgen Receptor Expression by Immunohistochemistry in Prostate, Possible Role in Diagnosis and Therapy.

Background: Recent discovery of gene rearrangements have brought a new look to the molecular pathogenesis of cancer. Gene fusions occur in nearly 60% of prostate adenocarcinoma, being the TMPRSS2-ERG one of the most common. Evidence supports the role of ERG fusion in tumorigenesis, progression and invasion via effecting pathways such as WNT, MYC, uPA, PI3K/AKT/PTEN, RAS/RAF/MAPF, NKX3.1, GST-pi and androgen receptor (AR) mediated signaling. Most of the ERG fusions involve 5'-partners androgen responsive. Therefore, we aimed to evaluate AR and ERG fusion protein expression on prostate tissue to find clinicopathological applications and possible role in therapy. Methods: One hundred three samples, including prostate core biopsies and radical prostatectomy specimens, were evaluated for ERG and AR expression by immunohistochemistry (IHC). ERG rearrangement was done by fluorescence in situ hybridization (FISH) on 11 randomly selected cases and correlated with IHC results. Results: From the total of 103 samples, eight (8/103) were benign, fourteen (14/103) had atypical glands, two (2/103) had prostatic intraepithelial neoplasia (PIN), and seventy nine (79/103) showed prostate adenocarcinoma. Forty four (44/79) tumor cases were Gleason score (GS) 6-7 (lower GS), and thirty five (35/79) were GS of 8-10 (higher GS). ERG immunoreaction was observed in 27.8% (22/79) of the tumor cases, showing higher expression in those with lower GS (68.2%, 15/22) compared to higher GS (31.8%, 7/22). Neither benign glands nor PIN stained with ERG. AR expression was observed in 75% of benign samples, 78.5% of atypical glands, 100% of PIN, and in 87.3% of tumor cases with no significant difference based on GS. Co-expression of ERG and AR was evaluated on all the tumor samples. ERG+/AR+ was seen in 77.3% (17/22) of the ERG+ tumor cases, with higher frequency in lower GS (64.7%, 11/17) compared to those with higher GS (35.3%, 6/17). All but five corresponding ERG+ tumor samples were negative for AR. Only 5 samples were ERG-/AR- corresponding to adenocarcinoma GS of 6. Presence or absence of ERG rearrangement was confirmed by FISH and correlated with IHC results. Conclusions: Characterization of ERG status by IHC in prostate tissue has an excellent correlation with FISH. It may also assist in diagnosis since none of the benign glands stained with ERG. Co-expression of ERG+/AR+ in prostate tumor by IHC may suggest gene fusion between ERG and a 5'-partner driven by androgen signaling such as TMPRSS2, which it could represent an important ancillary test for clinical management and development of new therapeutic targets.

CDK4 Amplification Reduces Sensitivity to CDK4/6 Inhibition in Fusion-Positive Rhabdomyosarcoma.

PURPOSE: Rhabdomyosarcoma (RMS) is the most common pediatric soft tissue sarcoma and includes a PAX3- or PAX7-FOXO1 fusion-positive subtype. Amplification of chromosomal region 12q13-q14, which contains the CDK4 proto-oncogene, was identified in an aggressive subset of fusion-positive RMS. CDK4/6 inhibitors have antiproliferative activity in CDK4-amplified liposarcoma and neuroblastoma, suggesting CDK4/6 inhibition as a potential therapeutic strategy in fusion-positive RMS. EXPERIMENTAL DESIGN: We examined the biologic consequences of CDK4 knockdown, CDK4 overexpression, and pharmacologic CDK4/6 inhibition by LEE011 in fusion-positive RMS cell lines and xenografts. RESULTS: Knockdown of CDK4 abrogated proliferation and transformation of 12q13-14-amplified and nonamplified fusion-positive RMS cells via G1-phase cell-cycle arrest. This arrest was mediated by reduced RB phosphorylation and E2F-responsive gene expression. Significant differences in E2F target expression, cell-cycle distribution, proliferation, or transformation were not observed in RMS cells overexpressing CDK4. Treatment with LEE011 phenocopied CDK4 knockdown, decreasing viability, RB phosphorylation, and E2F-responsive gene expression and inducing G1-phase cell-cycle arrest. Although all fusion-positive cell lines showed sensitivity to CDK4/6 inhibition, there was diminished sensitivity associated with CDK4 amplification and overexpression. This variable responsiveness to LEE011 was recapitulated in xenograft models of CDK4-amplified and nonamplified fusion-positive RMS. CONCLUSIONS: Our data demonstrate that CDK4 is necessary but overexpression is not sufficient for RB-E2F-mediated G1-phase cell-cycle progression, proliferation, and transformation in fusion-positive RMS. Our studies indicate that LEE011 is active in the setting of fusion-positive RMS and suggest that low CDK4-expressing fusion-positive tumors may be particularly susceptible to CDK4/6 inhibition.

Multifocal dysembryoplastic neuroepithelial tumours associated with refractory epilepsy.

Dysembryoplastic neuroepithelial tumours (DNET) are a common cause of tumour-associated epilepsy, and are usually located in the temporal lobes. We present a case of multifocal DNETs in both infra- and supra-tentorial locations, in a 23-year-old man with a coincident Type I Chiari malformation, presenting with medically refractory focal seizures. The extensive anatomical distribution of the lesions suggests a genetic component in their tumourigenesis.