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Supplemental Figure 1. Metformin effects on insulin, AMPK, mTOR

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Obesity is associated with risk of colorectal adenoma (CRA) and colorectal cancer. The signaling pathway activated by metformin (LKB1/AMPK/mTOR) is implicated in tumor suppression in ApcMin/+ mice via metformin-induced reduction in polyp burden, increased ratio of pAMPK/AMPK, decreased pmTOR/mTOR ratio, and decreased pS6Ser235/S6Ser235 ratio in polyps. We hypothesized that metformin would affect colorectal tissue S6Ser235 among obese patients with recent history of CRA. A phase IIa clinical biomarker trial was conducted via the U.S. National Cancer Institute-Chemoprevention Consortium. Nondiabetic, obese subjects (BMI ≥30) ages 35 to 80 with recent history of CRA were included. Subjects received 12 weeks of oral metformin 1,000 mg twice every day. Rectal mucosa biopsies were obtained at baseline and end-of-treatment (EOT) endoscopy. Tissue S6Ser235 and Ki-67 immunostaining were analyzed in a blinded fashion using Histo score (Hscore) analysis. Among 32 eligible subjects, the mean baseline BMI was 34.9. Comparing EOT to baseline tissue S6Ser235 by IHC, no significant differences were observed. Mean (SD) Hscore at baseline was 1.1 (0.57) and 1.1 (0.51) at EOT; median Hscore change was 0.034 (P = 0.77). Similarly, Ki-67 levels were unaffected by the intervention. The adverse events were consistent with metformin's known side-effect profile. Among obese patients with CRA, 12 weeks of oral metformin does not reduce rectal mucosa pS6 or Ki-67 levels. Further research is needed to determine what effects metformin has on the target tissue of origin as metformin continues to be pursued as a colorectal cancer chemopreventive agent.

BackgroundEarly evaluations of healthcare professional (HCP) COVID-19 risk occurred during insufficient personal protective equipment and disproportionate testing, contributing to perceptions of high patient-care related HCP risk. We evaluated HCP COVID-19 seropositivity after accounting for community factors and coworker outbreaks.MethodsPrior to universal masking, we conducted a single-center retrospective cohort plus cross-sectional study. All HCP (1) seen by Occupational Health for COVID-like symptoms (regardless of test result) or assigned to (2) dedicated COVID-19 units, (3) units with a COVID-19 HCP outbreak, or (4) control units from 01/01/2020 to 04/15/2020 were offered serologic testing by an FDA-authorized assay plus a research assay against 67 respiratory viruses, including 11 SARS-CoV-2 antigens. Multivariable models assessed the association of demographics, job role, comorbidities, care of a COVID-19 patient, and geocoded socioeconomic status with positive serology.ResultsOf 654 participants, 87 (13.3%) were seropositive; among these 60.8% (N = 52) had never cared for a COVID-19 patient. Being male (OR 1.79, CI 1.05–3.04,p = 0.03), working in a unit with a HCP-outbreak unit (OR 2.21, CI 1.28–3.81,p p = 0.05), and ethnically Latino (OR 2.10, CI 1.12–3.96,p = 0.02) were positively-associated with COVID-19 seropositivity, while working in dedicated COVID-19 units was negatively-associated (OR 0.53, CI = 0.30–0.94,p = 0.03). The research assay identified 25 additional seropositive individuals (78 [12%] vs. 53 [8%],p ConclusionsPrior to universal masking, HCP COVID-19 risk was dominated by workplace and community exposures while working in a dedicated COVID-19 unit was protective, suggesting that infection prevention protocols prevent patient-to-HCP transmission.Article summaryPrior to universal masking, HCP COVID-19 risk was dominated by workplace and community exposures while working in a dedicated COVID-19 unit was protective, suggesting that infection prevention protocols prevent patient-to-HCP transmission.

Primary bone marrow diffuse large B-cell lymphoma is an exceedingly rare form of non-Hodgkin lymphoma. It may demonstrate a leukemic presentation, and a proportion of cases have CD5 expression. The prognostic implications of this CD5-positivity remain unknown. Here, we present a 78-year-old man who presented with circulating peripheral blood lymphoma cells and a hypercellular marrow involved by diffuse large B-cell lymphoma, germinal center B-cell subtype. The patient responded favorably to six cycles of etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin, and rituximab (EPOCH-R) and intrathecal methotrexate. He unfortunately relapsed in several enlarged inguinal lymph nodes and succumbed to the lymphoma approximately one year after diagnosis, demonstrating the particularly aggressive clinical course of his disease.

Background: Early evaluations of healthcare professional (HCP) COVID-19 risk occurred during insufficient personal protective equipment and disproportionate testing, contributing to perceptions of high patient-care related HCP risk. We evaluated HCP COVID-19 seropositivity after accounting for community factors and coworker outbreaks.Methods: Prior to universal masking, we conducted a single-center retrospective cohort plus cross-sectional study. All HCP 1) seen by Occupational Health for COVID-like symptoms (regardless of test result) or assigned to 2) dedicated COVID-19 units, 3) units with a COVID-19 HCP outbreak, or 4) control units from 01/01/2020-04/15/2020 were offered serologic testing by an FDA-authorized assay plus a research assay against 67 respiratory viruses, including 11 SARS-CoV-2 antigens. Multivariable models assessed the association of demographics, job role, comorbidities, care of a COVID-19 patient, and geocoded socioeconomic status with positive serology.Results: Of 654 participants, 87 (13.3%) were seropositive; among these 60.8% (N=52) had never cared for a COVID-19 patient. Being male (OR 1.79, CI 1.05-3.04, p=0.03), working in a HCP-outbreak unit (OR 2.21, CI 1.28-3.81, pConclusions: Prior to universal masking, HCP COVID-19 risk was dominated by workplace and community exposures while working in a dedicated COVID-19 unit was protective, suggesting that infection prevention protocols prevent patient-to-HCP transmission.

Follicular lymphoma, the second most common non-Hodgkin lymphoma (NHL), primarily affects adults and shows an indolent clinical course. Rare cases of follicular lymphoma transform to a high-grade B-cell lymphoma with MYC and BCL2 rearrangements or “double-hit lymphoma”. Transformation to a “double-hit lymphoma” portends a worse prognosis and requires aggressive treatment. We report a comprehensive clinical, pathologic and radiographic review of a patient with previously undiagnosed low-grade follicular lymphoma that transformed into a “double-hit lymphoma”. The patient presented with a large heterogeneous mass 16 x 19 cm involving pancreatic head and neck and a mildly enlarged inguinal lymph node. Positron emission tomography (PET) study demonstrated Fluorodeoxyglucose (18F) (FDG)-avid peripancreatic mass. Tissue biopsy demonstrated a high-grade B-cell lymphoma with rearrangements t(14;18) and MYC, leading to the diagnosis of high-grade B-cell lymphoma with MYC and BCL2 rearrangements. Excisional biopsy of an inguinal lymph node demonstrated low-grade follicular lymphoma. Clonality studies demonstrated the same immunoglobulin clone V7-4 in inguinal lymph node and peripancreatic mass. Therefore, diagnosis of a high-grade B-cell lymphoma with MYC and BCL2 rearrangements that transformed from a low-grade follicular lymphoma was rendered. It is ultimately important to establish a tissue-based diagnosis at the different sites that are involved with lymphoma. Patient proceeded with the aggressive treatment with dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin and rituximab (EPOCH-R) treatment.

Background: Prodromal/pre-leukemic acute leukemia is rare, mostly described in childhood B-lymphoblastic leukemia (B-ALL), with its initial presentation confused with aplastic anemia (AA). Findings reported in pediatric prodromal/pre-leukemic B-ALL include female preponderance, bone marrow (BM) fibrosis and uniform hypocellularity, and transient recovery. Reports in adults are rarer still. We conducted a retrospective review of adult patients we diagnosed with B-ALL and mixed phenotype acute leukemia with B-lymphoid and myeloid components (MPAL-B/myeloid) with prodromal/pre-leukemic manifestations. We sought to identify common clinical features, peripheral blood (PB) and BM morphology, diagnostic considerations, and clinical follow-up (whenever available) to aid in improved recognition in the adult setting. Methods: Patients with unusual presentations preceding overt acute leukemia were identified from pathology and clinical records from our institution from 2010-2020. These include (a) incidental PB blasts and partial BM-based disease, followed by spontaneous, transient regression (STR); (b) pancytopenia with rare to absent PB blasts and unexpected detection of BM-based blasts (10% or lower), compatible with aleukemic prodrome (AP) cases; (c) pancytopenia without PB blasts yet unexpected excess BM-based blasts (50% or greater) followed by marked reduction of BM-based blasts (compared to initial BM) despite absence of treatment (AP-->SR). BM pathology during prodromal/pre-leukemic phases were reviewed to identify key morphologic features. Results: Seven patients were identified (4 M, 3 F; age range 20-75 years), including one previously described by our group (Table 1). All were of Hispanic ethnicity. They displayed a broad range of symptoms including fatigue, fevers, throat pain, and weight loss. Initial presentations were STR in 3, AP in 2, and AP-->SR in 2. Although variably cellular, all BM trephine biopsies in prodromal/pre-leukemic phase had at least focal hypocellularity with stromal damage, characterized by granular to fibrillary stroma, striking cellular dropout, and multiloculated fat cells with fibrinoid appearance. Relative erythroid hyperplasia was seen in 4, while dyserythropoiesis and dysmegakaryocytopoiesis were both observed in 2 cases, raising concern for myelodysplastic syndrome (MDS). AA and MDS were, in some cases, seriously considered in the initial differential diagnosis. Because of low level BM blasts at initial presentation in most (5/7), definitive lineage designation was sometimes challenging. Furthermore, flow cytometric data was limited and IHC stained only a sparse number of immature B-cells; thus normal precursor B-cell (hematogone) hyperplasia was a strong possibility. Workup in 2 cases during low numbers of BM-based blasts or spontaneous blast reduction (cases #6 and #7, respectively) revealed T-cell clones on PCR/NGS testing. In STR and AP cases, the disease proceeded from initial presentation to overt leukemia within a fairly short interval (range 1.5 - 7.0 months; median 2.5). Final diagnosis in the majority of cases (6/7) was B-ALL, with one case of MPAL-B/myeloid. Conclusion: Herein we describe a series of 7 adult prodromal/pre-leukemic cases of B-lymphoblasts (6 B-ALL; 1 MPAL-B/myeloid) with a variety of clinical presentations. In contrast to the female-predominant pediatric prodromal B-ALLs, most of our adult cases occurred in males. Given pancytopenia, relatively low numbers, and difficulty in assigning blast lineage, diagnostic considerations initially included MDS, AA, and hematogone hyperplasia with marrow regeneration. Key morphologic features that favored a prodromal/pre-leukemic case of B-lymphoblasts over MDS or AA were profound stromal damage and expansion (albeit low numbers) of immature B-cells - findings that would be unusual in low-grade MDS and AA. Intriguingly, in 2 of our cases, T-cell clones detected on PCR testing during low blast levels may indicate an immunologic role of T-cells in transiently reducing blasts in prodromal acute leukemia, as previously suggested by Zimmermannova and colleagues (Haematol2017;102:e225-228). Regardless of whether patients experienced STR, AP, or AP-->SR, close clinical follow-up is advised, as progression to overt leukemia occurs within a relatively short time frame (median of 2.5 months in our series). Disclosures No relevant conflicts of interest to declare.

Background Chronic lymphocytic leukemia (CLL) is a clonal disorder of B lymphocytes, characterized by proliferation of small mature lymphocytes involving the blood, bone marrow, and lymphoid tissues. CLL can rarely involve the central nervous system (CNS), either by involving brain parenchyma or cerebrospinal fluid (CSF). We present a series of three cases with clinically significant involvement of the CNS with CLL. Results During the past 2 years, our medical center managed three patients with CLL who presented with symptomatic CNS involvement, as determined by flow cytometry. The immunophenotypic profile was that of a typical CLL with light chain restricted small B cells positive for CD20 (dim) and coexpressing CD5 and CD23. In addition, two patients had brain involvement by SLL that was confirmed by brain biopsy. Notably, the brain lesions had a mildly elevated Ki-67 proliferative index (10%-30%). Bone marrow was involved in two patients, showing nodular, interstitial, and diffuse patterns. Bone marrow involvement ranged from 60% to 80% and showed very low Ki-67 proliferative index. None of the patients had features suggestive of a CLL transformation. FISH was performed on either bone marrow or CSF and demonstrated that patient 1 had Del11q(ATM) and Dell13q, patient 2 had trisomy 12, and patient 3 had del17(TP53) and IGH rearrangement. All of the patients showed persistent CSF involvement, ranging from 4 to 12 weeks, requiring aggressive treatment with intrathecal chemotherapy. At the end of treatment, all of the patients were clear of CNS involvement as judged by flow cytometry of CSF. Conclusion We report three patients who had persistent involvement of CSF. Only one patient had del17(TP53), a cytogenetic feature that is associated with high-risk CLL. It would be interesting to study clonal evolution of CLL to understand the mechanisms that underlie involvement of the CNS.

ContextLymphoid aggregates are seen in a minority of bone marrow biopsy specimens, and when present, their neoplastic nature is often apparent by morphologic evaluation. However, the distinction between benign and malignant aggregates can be a diagnostic challenge when there are multiple aggregates with no documented history of lymphoma.ObjectiveTo aid in the distinction between benign and malignant B-cell lymphoid aggregates.DesignPreviously, we described specific distribution patterns for B and T lymphocytes within bone marrow aggregates. To statistically analyze the significance of these patterns as well as previously reported criteria, we examined 128 bone marrow specimens with benign aggregates and 78 specimens with documented malignant B-cell aggregates and calculated specific odds ratios (ORs) and 95% confidence intervals (CIs) to aid in differentiating between benign and malignant B-cell aggregates.ResultsAggregates with infiltrative edges (OR, 80.54; 95% CI, 31.76–204.21), a B-cell pattern (OR, 30.08; 95% CI, 13.28–68.10), paratrabecular location (OR, 10.17; 95% CI, 3.96–26.12), size greater than 600 μm (OR, 6.83: 95% CI, 3.61–12.93), or cytologic atypia correlated with malignancy.ConclusionWhen taken collectively, the presence of more than 2 of these characteristic features was strongly predictive of malignancy.

Author(s): Farzaneh, Ted; Wang, Beverly; Clark, Nancy; Eade-Viele, Carol; Kelner, Joyce; Rodgers, Jennifer; Sempa, Geoffrey; Detweiler, Neil; Kasmarek, Tara; Lee, Thomas; Zhao, Xiaohui; Monuki, Edwin S; Rezk, Sherif A | Abstract: BackgroundTraining in patient safety, quality, and management is a key component of Graduate Medical Education (GME) training in all specialties. However, residency programs, especially Pathology programs, often find it challenging to create strong learning opportunities in these areas.ObjectivesFocused quality assurance (QA) projects are one approach to teach and engage trainees in these key areas. Residents have been historically involved in different QA projects in our department but mainly in small secondary roles. Leading a large QA project that can enhance residents' management skills and improve clinical operations in our laboratory was the main objective of our project.DescriptionA new process for laboratory self-inspection led by residents was implemented that simulates the exact process of a formal outside College of American Pathologists (CAP) inspection. We aim to prove that resident-led QA activities not only have profound educational benefit but can also result in significant performance and operational improvement.ResultsFor this paper, we focus on the Histology laboratory since the ramifications from the self-inspection process during a three year period were profound leading to change in management, workflow changes, and notable improvement in staff morale.ConclusionThe self-inspection process exposed the residents to operational issues and corrective actions that provided them the opportunity to take a more active role in laboratory management and helped prepare them for post-graduation challenges. It also helped the department identify and rectify many operational issues, confirmed by the enumeration of CAP deficiencies and significant improvement of staff morale.

Objectives: To report a patient with primary effusion lymphoma who was negative for human herpesvirus-8 (HHV-8), human immunodeficiency virus, Epstein-Barr virus, hepatitis C virus, and hepatitis B virus, as well as review 54 reported cases of HHV-8–unrelated primary effusion lymphoma (PEL)–like lymphoma in the literature to clarify the nature of this entity. Methods: The patients’ characteristics, clinical presentation, pathogenesis, morphologic-immunophenotypic features, clinical management, and prognosis were studied. Results: HHV-8–negative PEL-like lymphomas often occur in immunocompetent and elderly patients, are sometimes associated with chronic inflammation–related fluid overload, are mostly large B-cell or large B-cell with plasmacytic differentiation type, and are associated with a better prognosis. Conclusions: In various aspects, HHV-8–unrelated PEL-like lymphoma is a different entity from HHV-8–related PEL. Immunophenotype, morphology, and c-myc/8q24 status should be included for differential diagnosis. A test for c-myc or 8q24 abnormalities should be recommended for subdividing HHV-8–unrelated PEL-like lymphoma, which may have benefits in patient management.

Indeterminate cell tumor (ICT) is a rare neoplastic dendritic cell disorder that has been poorly defined due to its rarity and poorly understood histogenesis and pathogenesis. It is characterized by a proliferation of dendritic cells, which mimic Langerhans cells immunophenotypically (positive for CD1a and S-100 protein), but lack Birbeck granules characteristic of Langerhans cells. The clinical, morphologic, immunophenotypic, and ultrastructural features of 5 ICT cases are reported in an attempt to further define ICT and to examine the postulated relationship between indeterminate cells and Langerhans cells. Four of 5 patients were females, and 4 of 5 were older than 68 years. Three of 5 patients had cutaneous lesions, whereas 2 presented with cervical lymph node involvement. Two patients had a possible association with lymphoma: first patient had a history of progressive follicular lymphoma that led to patient's demise and the second patient had unexplained systemic lymphadenopathy and died 1 week after the biopsy. All 5 ICT cases expressed CD1a and S-100 protein, but lacked Langerin expression and Birbeck granules ultrastructurally. Interestingly, a t(14;18) was detected by fluorescence in situ hybridization in the ICT cells of the patient with previous follicular lymphoma and a monoclonal κ light chain gene rearrangement was detected by polymerase chain reaction in the patient with systemic lymphadenopathy. In both cases, there was no morphologic or immunophenotypic evidence of a concurrent B-cell lymphoma. In conclusion, ICT is a rare neoplasm that can occur de novo or in association with a B-cell lymphoma, possibly as a result of B-cell dedifferentiation caused by relatively unknown mechanisms. Finally, Langerin immunostaining may be used as a surrogate marker for the ultrastructural demonstration of Birbeck granules, the absence of which represents a strong diagnostic criterion for ICT. © 2008 Lippincott Williams & Wilkins.

Meckel diverticulum is the most common congenital defect of the gastrointestinal tract. It can be asymptomatic or mimic appendicitis and may be complicated by bleeding, diverticulitis, obstruction, and, rarely, neoplasia. We report the first case of extranodal marginal zone lymphoma occupying a Meckel diverticulum. A 44-year-old man with history of colonic diverticulitis presented to the emergency department for evaluation of acute abdominal pain. Radiography showed enteric obstruction, prompting diagnostic laparoscopy. Above the level of mid-ileum an intact Meckel diverticulum was identified. Microscopy showed extensive infiltration of sheets of small lymphocytes with abundant cytoplasm (monocytoid B-cells) prominently in submucosa and focally transmural involving serosal adipose tissue with multiple reactive germinal centers. The immunostains showed positivity for CD20, BCL-2, and CD43 (weak) and negativity for CD3, CD5, BCL-1, CD10, and BCL-6 in monocytoid B-cells. Fluorescence in situ hybridization studies revealed API2-MALT1 fusion signals consistent with t(11;18)(q21;q21), which confirmed the diagnosis of extranodal marginal zone lymphoma, also known as mucosa associated lymphoid tissue lymphoma.

Benign lymphoid aggregates are seen in only a minority of bone marrow specimens, but their distinction from non-Hodgkin lymphoma, particularly B-cell lymphomas, can represent a diagnostic challenge. Although criteria have been proposed to help distinguish between benign and malignant aggregates, a detailed description of the distribution patterns of B and T lymphocytes within benign lymphoid aggregates has not been investigated. One hundred thirty-seven cases of bone marrow specimens containing benign aggregates were studied with a panel of immunostains. A subset of these cases was also examined for immunoglobulin gene rearrangements by polymerase chain reaction. The aggregates were categorized based on size, location (paratrabecular or random), presence of infiltrating edges, and distribution of lymphoid cell populations. In addition, we examined 40 cases of bone marrow biopsies with documented malignant lymphoid aggregates for comparison purposes. We report that the distribution of B and T lymphocytes within lymphoid aggregates may serve as a useful criterion to aid in the separation between benign and malignant aggregates. When aggregates exhibit a predominance of T cells, consist of a central core of T cells surrounded by a rim of B cells, or have a mixed distribution of B and T cells, they are more likely to be benign. On the other hand, an increased likelihood of malignancy occurs when aggregates exhibit a predominance of B cells or consist of a central core of B cells surrounded by a rim of T cells (excluding germinal center formation), and assessing other features worrisome of malignancy (large aggregate size, presence of infiltrative edges, cellular atypia, and paratrabecular location, among others) is warranted. © 2013 Elsevier Inc.

Solitary fibrous tumor (SFT) is an uncommon spindle cell neoplasm of increasing incidence that was originally described to be of pleural origin; however, more recently, SFT has been reported in extrapleural sites, including the orbit, liver, salivary glands, tongue, nose, paranasal sinuses, larynx, retroperitoneum, meninges, and thyroid. The increase in the number of SFTs does not necessarily mean increased incidence of this tumor but rather an increased understanding of this tumor, especially recognition of this tumor in extrapleural locations, which has been aided by immunohistochemical analysis. We report a case of SFT in the auditory canal, which to our knowledge has not been previously reported, as evident by morphologic findings and immunophenotype.

Insertion or deletion mutations in calreticulin (CALR) are present in the majority of JAK2V617F-negative MPN patients. We utilized a murine retroviral transduction-transplantation model to express the 52bp CALR deletion mutation (CALRDEL) in both BALB/c and C57B/6 backgrounds. As described previously (Marty et al., Blood 2016;127:1317), recipients of CALRDEL-transduced marrow developed persistent thrombocytosis without leukocytosis or erythrocytosis by two months post-transplant. Mice were euthanized at six and nine months post-transplant to evaluate the tempo of disease progression. At six months CALRDEL mice had impressive expansion of megakaryocytes expressing the CALRDEL mutant protein in the bone marrow (BM) without fibrosis or significant splenomegaly. By nine months BM fibrosis and splenomegaly were present. Both whole BM and spleen cells were able to serially transplant the MPN phenotype into secondary recipients. When cultured in collagen-based media supplemented with thrombopoietin, CALRDEL BM cells produced an increased number of megakaryocyte colonies as compared to empty vector. The increased colony formation potential of CALRDEL bone marrow cells was limited to megakaryocytes, we found no increase in colony formation from CALRDEL hematopoietic stem and progenitor cells in methylcellulose with cytokines supporting erythroid and GM colony formation. However, CALRDEL enhanced the serial replating ability of LKS (lineageneg, c-kit+ Sca-1+) cells. Both pSTAT5 and pERK were increased in whole spleen lysates from CALRDEL mice as compared to wild-type BALB/c mice. Therefore, we tested the impact of ruxolitinib, a JAK1/2 inhibitor, and trametinib, a MAPK/ERK inhibitor, on the MPN phenotype of CALRDEL mice. At six months post-transplant mice were treated with either ruxolitinib (90mg/kg PO BID), trametinib (3mg/kg PO daily), or vehicle for 40 days. Ruxolitinib reduced pSTAT5 but caused a paradoxical increase in pERK in whole spleen lysates, while trametinib reduced pERK but not pSTAT5. Trametinib caused a transient increase in platelets and white cells. In spite of pharmacodynamic evidence of effective dosing, ruxolitinib had no significant effect on platelet or leukocyte count but did reduce hemoglobin slightly. Both ruxolitinib and trametinib reduced spleen weight. Ruxolitinib reduced the fraction of the mutant CALRDEL allele (inferred from percentage of GFP+ cells) in the spleen but not the bone marrow, while trametinib had no impact on disease allele burden in any organ. Neither ruxolitinib nor trametinib reduced the expansion of megakaryocytes in the bone marrow but trametinib significantly reduced marrow fibrosis (average score MF-2.5 for vehicle, MF-1.75 for ruxolitinib, MF-1 for trametinib). To assess the role of STAT5 in the pathogenesis of the ET-like MPN induced by the CALRDEL mutant, we transduced BM from syngeneic Balb/c donors carrying a floxed Stat5ab allele in combination with a Stat5ab null allele (Mx-Cre;Stat5abfl/-; Walz et al., Blood 2012;119:3550). Haploinsufficiency for Stat5ab significantly delayed the development of ET-like MPN and attenuated thrombocytosis, implicating JAK2-STAT5 signaling directly in the pathogenesis of this disease. In summary, this CALRDELmouse model results in an MPN phenotype resembling essential thrombocythemia followed by myelofibrosis. CALRDELresults in expansion of megakaryocytes and platelets without expansion of other myeloid cell types. Both pSTAT5 and pERK are increased in our CALRDEL model and pharmacologic inhibition of pERK results in reduction of fibrosis without reducing megakaryocytes. These studies implicate pERK as a potential anti-fibrosis therapeutic target in MPN. Disclosures No relevant conflicts of interest to declare.

Correspondence ackn owledgements British Lung Foundation UK and Lung Pathology Departmental Funds for funding. M Tsoumakidou J Zhu Z Wang A Thorley 1 S Kemp 1 T Tetley 1 P K Jeffery Lung Pathology, Department of Gene Therapy, and Lung Cell Biology, Section of Airways Disease, National Heart & Lung Institute, Imperial College London, London, UK 1. Vermaelen K, Pauwels R. Pulmonary dendritic cells. Am. J. Respir. Crit. Care Med. 2005; 172; 530–551. 2. Demedts I, Brusselle G, Vermaelen K et al. Identification and characterization of human pulmonary dendritic cells. Am. J. Respir. Cell Mol. Biol. 2005; 32; 177–184. 3. Tsoumakidou M, Tzanakis N, Papadaki HA et al. Isolation of myeloid and plasmacytoid dendritic cells from human broncho- alveolar lavage fluid. Immunol. Cell Biol. 2006; 84; 267–273. 4. Todate A, Chida K, Suda T et al. Increased numbers of dendritic cells in the bronchiolar tissues of diffuse panbronchiolitis. Am. J. Respir. Crit. Care Med. 2000; 162; 148–153. 5. Witherden IR, Vanden Bon EJ, Goldstraw P et al. Primary human alveolar type II epithelial cell chemokine release: effects of cigarette smoke and neutrophil elastase. Am. J. Respir. Cell Mol. Biol. 2004; fixation. The metal suture is later trimmed off, allowing perpendicular slices to be made across the lung ⁄ pleural tissue, permitting detailed analysis. One issue not covered in any detail in the review is that of occupational dust-related pathology and emphysema. Assessment of emphysema is particularly important at autopsy in this regard, as it is used to substantiate ⁄ refute medicolegal claims by relatives of the deceased. It is vital to try to inflate postmortem lung tissue (in a similar manner) for ideal quantification of disease and, even if whole mount lung tissue sections are not available, small, adequately inflated lung samples can produce significant information that can be tested in the legal setting. In addition, it is worth considering other occupa- tional disease, particularly in relation to metal fumes. Cadmium exposure (for example) can produce signifi- cant emphysematous change many years after expo- sure. 2 Retention of some lung tissue for mass spectroscopy can be particularly useful if one is considering metal or other chemical-related disease. Finally, whilst I would entirely concur with the pathological approach to classification given in the review, I would suggest that any analysis of lung parenchyma must be cross-correlated against respira- tory function tests and other clinical information. Of particular value is computerized tomography ⁄ magnetic resonance imaging in terms of extrapolation of small biopsy ⁄ autopsy sample histology in order to derive full appreciation of overall pulmonary pathology status. 3 S K Suvarna Advances in the pathology of COPD Department of Histopathology, Northern General Hospital, Sheffield, UK DOI: 10.1111/j.1365-2559.2007.02816.x Sir: I read with great interest the very detailed and helpful review on emphysema, 1 and agree this is the gold standard approach to assessment of lung tissue for chronic obstructive pulmonary disease (COPD). Regret- tably, autopsy and tissue retention issues in the UK have led to problems with examining lung parenchyma in this format and in most cases only small samples of tissue can be retained for COPD assessment. For surgical samples, often submitted as thoraco- scopic (video-assisted thorascopic surgery) samples with wire suture material at the margins, I would recommend inflation of these samples upon receipt in the laboratory by injecting formalin (by syringe ⁄ needle) into the lung parenchyma through the pleura, until the sample looks adequately inflated for ideal 1. Wright JL, Churg A. Advances in the pathology of COPD. Histopathology 2006; 49; 1–9. 2. Churg A, Colby TV. Diseases caused by metals and related com- pounds. In Churg A, Green FHY eds. Pathology of occupational lung disease, 2nd edn, Baltimore: Williams and Wilkins, 1998; 77–128. 3. Muller NL, Fraser RS, Lee KS, Jokhoh T. Diseases of the lung. Radiological and pathological correlations. Philadelphia: Lippincott Williams & Wilkins, 2003; 239–254. Myofibroblastoma of the male breast: a rare entity of increasing frequency that can be diagnosed on needle core biopsy DOI: 10.1111/j.1365-2559.2007.02808.x Sir: Myofibroblastoma is a rare benign mesenchymal tumour that was first described by Wargotz et al. in 2007 The Authors. Journal compilation 2007 Blackwell Publishing Ltd, Histopathology, 51, 552–580.

Epstein-Barr virus (EBV) is a member of the human herpesvirus family that was initially isolated from a cultured Burkitt lymphoma cell line by Epstein et al in 1964. Subsequent studies have proven that it is the causative agent in most cases of infectious mononucleosis. Primary infection is usually asymptomatic in childhood; but in adulthood, it is associated with a self-limiting infectious mononucleosis syndrome in approximately one third of the cases. EBV has been linked to many human neoplasms including hematopoietic, epithelial, and mesenchymal tumors. In this review, we will only discuss the EBV-associated lymphoproliferative disorders, dividing them into B-cell, T/NK-cell, and HIV-related lymphoproliferative disorders.

Introduction: Chronic natural killer (NK) cell lymphoproliferative disorders comprise a rare subgroup of diseases characterized by proliferation of NK cells with expression of CD3-/CD16+ or CD56+ on flow cytometry. Clinically, these patients present with cytopenias as well as fatigue and B symptoms. The diagnosis can be challenging due to frequent lack of a unique clonal marker. Chronic NK cell lymphoproliferative disorder can be differentiated from NK cell leukemia based on the absence of chronic EBV infection. Macrophage activation syndrome (MAS) can occur in the setting of hematologic malignancies. Due to the rarity of this diagnosis, there are no randomized controlled trials to determine the optimal therapy. We report the case of an elderly gentleman whom we diagnosed with chronic NK cell lymphoproliferative disorder, complicated by transfusion-dependent autoimmune hemolytic anemia, and MAS, and who was successfully treated with oral cyclophosphamide. Case Report: In March 2014, a 75 year old Caucasian male, who was otherwise in good health, had a syncopal episode. Two months earlier, he had reported dyspnea on exertion and cough. He was diagnosed with bronchitis and these symptoms resolved with a course of antibiotics. Cardiac work-up was unremarkable. Complete blood count demonstrated white blood count of 16.8K with 84% lymphocytes, with hemoglobin of 7.8g/dL and platelets of 264K. Lactate dehydrogenase (LDH) was elevated at 615 IU/L, haptoglobin was undetectable, total bilirubin was mildly elevated at 1.5mg/dL and a Coomb's assay was negative. The patient had adequate iron stores with 50% saturation and erythropoietin level was 76.7ug/L. CT of the abdomen/pelvis demonstrated mild splenomegaly of 14cm. CT scan also revealed numerous thoracic-spine soft lesions. Bone marrow biopsy revealed a hypercellular marrow with appropriate trilineage hematopoiesis, erythroid hyperplasia and increased lymphocytes which were CD2+ CD7+ CD16+ and CD 56 negative cells as well as negative for T and B cell markers. This lymphocyte population was identified as NK cells and comprised 16% of his cells. The patient's transfusion dependence did not decrease despite oral prednisone. He transferred to our university medical center. Repeat flow cytometry after one month on prednisone demonstrated an increase to 70% of the lymphocytes. Further, there was a high suspicion for macrophage activation syndrome given his ferritin of 7,358ng/mL, triglycerides of 290mg/dL and interleukin-2 receptor of 7,250pg/mL. There was no evidence of active hemophagocytosis on the bone marrow biopsy. He continued on prednisone 60mg daily, and started on cyclophosphamide 50mg daily. His clinical course over the next several months was complicated by recurrent fevers, night sweats, mental status changes, and hyponatremia. Sepsis was ruled out as a cause of his fevers and his mental status changes were attributed to be secondary to hyponatremia and MAS. Work-up of his hyponatremia revealed syndrome of inappropriate antidiuretic hormone secretion (SIADH). After 2 months of treatment, his sodium level and mental status normalized. His cyclophosphamide was increased to 100mg daily and maintained at that dose. He was then tapered off prednisone. This patient's B symptoms of fevers and night sweats abated after several months of this therapy, and his transfusion dependence and underlying MAS resolved. The patient has been maintained on cyclophosphamide 100mg for 11 months and has shown resolution of his lymphocytosis and normalization of his blood counts. Discussion: Chronic lymphoproliferative disorders of NK cells continue to be a difficult group of diseases to recognize. Moreover, this case was further complicated by MAS based on elevated interleukine-2 receptor, triglycerides and ferritin levels in the setting of B-symptoms and liver enzyme elevations. This constellation of factors has not been previously described in the literature. Due to lack of clonal markers in NK cells, the diagnosis is frequently made by exclusion. The patient has done very well on oral cyclophosphamide and prednisone alone. We present this case to increase provider awareness and hopefully allow for improved diagnosis and treatment options in the future. Disclosures No relevant conflicts of interest to declare.

Background: Despite advances in colorectal cancer (CRC) screening, early detection, and treatment, CRC remains the 2nd most common cancer cause of death in the U.S.. Obesity is increasing in incidence in the U.S., and has been implicated in colorectal adenoma (CRA) risk, risk of CRA recurrence, and risk of CRC. Obese patients with history of CRA are a high-risk group that may benefit from novel CRC prevention strategies. There is early evidence for reduced cancer mortality among metformin users. The signaling pathway activated by metformin (LKB1/AMPK/mTOR) is implicated in tumor suppression in ApcMin/+ mice, as evidenced by metformin-induced reduction in polyp burden, increased ratio of pAMPK/AMPk, decreased ratio of pmTOR/mTOR, and decreased ratio of pS6Ser235/S6Ser235 in polyp specimens. We hypothesized that metformin would affect colorectal tissue S6Ser235 similarly in humans, targeting obese patients with recent history of CRAs as a high-risk group. Methods: A phase IIa clinical biomarker trial was conducted across 3 clinical sites via the NCI-funded Southern California Chemoprevention Program. Eligible participants included non-diabetic, obese patients (BMI >30) with history of colorectal adenoma within the past 3 years, age ≥ 35 years and ≤ 80 years. All patients received an upward titration of metformin over 3 weeks to 1000mg po bid, which was continued until the end-of-study (EOS) at 12 weeks. Rectal mucosa biopsies were obtained at baseline (BL) and at time of EOS endoscopy. Tissue S6Ser235 immunostaining was analyzed in a blinded fashion by the study pathologist using Histo Score (HScore) analysis. A paired t-test was used to examine the effect of metformin on activated S6serine235 (i.e., the ratio of pS6serine235/ S6serine235). Results: 45 patients were consented to achieve 32 eligible subjects. 4 subjects were removed from study due to Adverse Events (1 SAE, unrelated). In order of frequency, the most common AEs were diarrhea, cramping, flatulence, nausea, stomach pain; 80% of participants had Grade 1 AE, 27% had grade 2 AE. Mean (SD) weight and body mass index at BL were 105.2 (17.42) kg and 34.9 (5.57) respectively. Weight did not significantly differ over the course of the study. Glucose levels at EOS did not significantly differ from BL. Vitamin B12 levels were significantly reduced at EOS vs. BL (-46.7 pg/mL, 95% CI -73.2 to -20.2). Comparing EOS to BL tissue S6Ser235 by IHC HScore analysis, no significant differences were observed. Mean (SD) Hscore at BL was 1.1 (0.57) and 1.1 (0.51) at EOS. Median HScore change was 0.032 (p=0.77). Conclusions: Among obese CRA patients, 12 weeks of oral metformin 1000mg twice daily does not reduce pS6 levels in the rectal mucosa. Other potential mechanisms of action have not yet been analyzed. Data from this clinical trial indicate that metformin can be used safely in a non-diabetic population. Further research is needed to determine what effects, if any, metformin has on the target tissue of origin (colorectum) relevant to colorectal carcinogenesis if metformin is to be pursued as a CRC chemopreventive agent. Citation Format: Jason A. Zell, Christine E. McLaren, Timothy R. Morgan, Michael J. Lawson, Sherif Rezk, Gregory C. Albers, Wen-Pin Chen, Joseph C. Carmichael, Luz Rodriguez, Eva Szabo, Leslie Ford, Michael Pollak, Frank L. Meyskens. A Phase IIa trial of metformin for colorectal cancer risk reduction among patients with a history of colorectal adenomas and elevated body mass index. [abstract]. In: Proceedings of the Thirteenth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2014 Sep 27-Oct 1; New Orleans, LA. Philadelphia (PA): AACR; Can Prev Res 2015;8(10 Suppl): Abstract nr A21.

Acute lymphoblastic leukemia (ALL) with eosinophilia is an uncommon disorder with several distinctive clinical and pathologic features. Eosinophilia associated with ALL usually precedes the diagnos...

DP-1 is a G1 cell cycle-related protein that forms heterodimers with E2F, a family of transcriptional factors regulating the expression of genes important for G1 to S progression. Although the exact role of DP-1 is not well understood, it has been shown to stabilize DNA binding of E2F proteins. By immunohistochemistry, the authors examined the expression of DP-1 in lymphoid tissues, including 8 cases of reactive follicular hyperplasia and 69 cases of B-cell non-Hodgkin lymphoma. The expression of the cell cycle-related proteins E2F-1 and Ki-67 was also assessed. Scoring was based on the proportion of labeled nuclei (1-10%, 11-25%, 26-50%, and > 50%). In reactive follicular hyperplasia, staining for DP-1, E2F-1, and Ki-67 was largely confined to the germinal centers. All 25 cases of follicular lymphoma, regardless of grade, had a high proportion (> 50%) of DP-1-positive cells but a lower proportion of cells marking for E2F-1 and Ki-67 (P < 0.001). The diffuse large B-cell lymphomas (n = 24) had high DP-1 and Ki-67 scores but low E2F-1 scores (P < 0.001). Small lymphocytic (n = 10), marginal zone (n = 3), and mantle cell lymphomas (n = 5) contained relatively low proportions of cells labeled for all three markers. Precursor B-cell lymphoblastic lymphoma (n = 2) displayed high proportions of cells positive for DP-1, Ki-67, and E2F-1 (> 50% in both cases). Except in follicular center cell lesions, DP-1 expression generally correlated with that of Ki-67. However, the expression of DP-1 was discordant with that of E2F-1 in benign and malignant follicular center cells, suggesting that DP-1 may have functions other than facilitating E2F-1-dependent gene regulation and cell cycle progression in these neoplasms.

Cyclin D1 and E2F-1 proteins are essential for the regulation of the G1/S transition through the cell cycle. Cyclin D1, a product of the bcl-1 gene, phosphorylates the retinoblastoma protein, releasing E2F-1, which in turn activates genes involved in DNA synthesis. Expression patterns of E2F-1 protein in thyroid proliferations have not been reported. This study used monoclonal antibodies for cyclin D1 and E2F-1 proteins to immunostain sections of normal thyroid, hyperplastic (cellular) nodules, follicular adenomas, follicular carcinomas, and papillary carcinomas. The proliferation rate was examined using an antibody specific for the Ki-67 antigen. Fluorescence in situ hybridization (FISH) methods and chromosome 11-specific probes were also employed to determine chromosome copy number and to assess for evidence of amplification at the 11q13 locus in papillary and follicular carcinomas with cyclin D1 overexpression. Concurrent overexpression of Ki-67, cyclin D1, and E2F-1 was found in the majority of benign and malignant thyroid lesions, compared with normal thyroid tissue. Cyclin D1 up-regulation was not due to extra copies of chromosome 11, or bcl-1 gene amplification. Malignant tumours showed the highest expression for all three markers, particularly papillary carcinomas. E2F-1 was detected at the same or slightly lower levels than cyclin D1. It was only found when cyclin D1 was overexpressed. Because cyclin D1 normally activates E2F-1, up-regulation of cyclin D1 may lead to E2F-1 overexpression in benign and malignant thyroid lesions.

The role of cell cycle protein expression in gestational trophoblastic disease is poorly understood. In this study we investigated the immunostaining patterns of G(1) restriction point and G(1)-S regulatory proteins E2F-1, Cdk2, cyclin E, p27(kip1), and the proliferation marker Ki-67 on routinely processed sections of 29 hydatidiform moles (10 partial moles and 19 complete moles, including 9 persistent moles), 7 choriocarcinomas, and 7 normal placentas. Ki-67 trophoblast staining decreased with increasing gestational age of the placenta, and showed maximal expression in gestational trophoblastic disease. Cyclin-dependent kinase activity, as reflected by Cdk2 expression patterns, also decreased with placental maturation. E2F-1 was uniquely expressed by trophoblasts of moles and choriocarcinoma. Cyclin E was maximally expressed by complete moles and choriocarcinomas, and showed an inverse relationship with the cyclin-dependent kinase inhibitor p27(kip1). Abnormal trophoblastic proliferations may be mediated through interactions of Cdk-2, E2F-1, cyclin E, and p27(kip1). Overexpression of cyclin E was associated with more aggressive forms of gestational trophoblastic disease. However, we did not find distinguishing features between complete moles that spontaneously resolved after evacuation and persistent moles that required chemotherapy. The different expression patterns of cyclin E and E2F-1 in partial and complete moles may be useful in distinguishing these two entities. Furthermore, loss of p27(kip1) in malignant trophoblast may represent a necessary step in the development of choriocarcinoma.

Plasma cell neoplasms (PCNs) refer to a group of true plasma cell disorders that include multiple myeloma, plasmacytoma, primary amyloidosis, and light and heavy-chain deposition diseases. Epstein-Barr virus (EBV) expression has been reported in several hematopoietic and nonhematopoietic disorders like Hodgkin lymphoma, Burkitt's lymphoma and nasopharyngeal carcinoma. Till recently, EBV expression in PCNs has been limited to cases with …

Solitary plasmacytoma of the bone (SPB) is a rare plasma cell neoplasm that usually presents as a lytic lesion mainly localized within the axial skeleton with no accompanying paraproteinemia or bone marrow involvement. The occurrence of SPB in young individuals is exceedingly rare. Trauma can cause enhanced release of cytokines resulting in increased proliferation of plasma cells. …