Multiple Myeloma Oncogene Research Articles

Diagnosis of canine B-cell chronic lymphoid leukemia with a CD21 negative phenotype using the LT21 clone CD21 antibody in flow cytometry: a case report

BackgroundChronic lymphoid leukemia (CLL) is a hematological disorder characterized by the clonal expansion of small mature lymphocytes that accumulate in the blood and bone marrow. CLL can arise from B-, T-, or natural killer cell clones. The cytological evaluation of blood smears is often the simplest and least invasive method for diagnosing lymphoid leukemia. Immunophenotyping is used to further subclassify the type of lymphoid leukemia.Case presentationA 15-year-old, 4.4-kg spayed female Shih Tzu was presented to the veterinary medical teaching hospital of Kangwon National University. Despite having a normal appetite and activity level, cervical and inguinal lymph node enlargement was noted on physical examination. Complete blood count revealed severe leukocytosis, severe lymphocytosis, and monocytosis. Splenomegaly, hepatomegaly, and lymph node enlargement were detected on radiographic and ultrasonographic examination. Immunophenotyping was performed using peripheral blood mononuclear cells (PBMCs). The majority of lymphocytes exhibited the following profiles: CD3−CD79a− (97.5%), CD4−CD8− (98.6%), CD21−CD79a− (98.4%), CD34− (0.1%), CD45+ (99.6%), major histocompatibility complex class II+ (99.5%), and CD14− (0.5%). Based on the immunophenotyping results, possible differentials considered included the following: the majority of lymphocytes may be natural killer (NK) cell clones, plasma cell clones, or show aberrant expression or loss of CD21 marker due to the neoplastic nature of the cells. Further flow cytometry was performed using antibodies against CD3, CD5, CD94, and granzyme B. The combined results indicated that the predominant lymphocyte subset in the PBMCs was CD3−CD5−CD21−CD94−granzyme B−. To confirm monoclonality and exclude the aberrant loss of CD markers, a polymerase chain reaction for antigen receptor rearrangement (PARR) assay was conducted. The PARR assay, using DNA from blood and lymph node samples, showed B-cell monoclonality. Immunocytochemistry using PBMCs showed that the plasma cell marker Multiple Myeloma Oncogene 1 (MUM1) was not expressed. Therefore, the diagnosis was confirmed to be B-cell CLL.ConclusionImmunophenotyping can help subclassify the type of lymphoid leukemia; however, as tumor cells can show aberrant expression or loss of the CD21 marker, combining immunophenotyping with the PARR assay could yield a more accurate diagnosis.

Clinical, Pathological and Immunohistochemical Characteristics of Patients with Primary Testicular Lymphoma Undergoing Orchiectomy: A Systematic Review and Meta-Analysis.

Primary testicular lymphoma (PTL) is a rare and aggressive malignant tumour with no specific clinical symptoms. Large-scale evidence-based medical evidence to guide preoperative diagnosis is lacking at present. This study aimed to analyse the clinical, pathological and immunohistochemical characteristics of patients with PTL undergoing testicular resection surgery. Literature on the clinical characteristics of patients with PTL undergoing orchiectomy was retrieved from databases, including PubMed, Web of Science, Embase, Cochrane Library, China National Knowledge Infrastructure (CNKI) and Wanfang Data. The search covered all available records from the inception of these databases until December 31, 2023. Data extraction was followed by a meta-analysis using Stata 15.0 software. A total of 22 articles and 475 cases of PTL were included. The meta-analysis revealed that 58.1% of patients with PTL undergoing orchiectomy were under 60 years old, and 41.9% were 60 years or older. The lesion is mostly located on the right side (55.1%). Common symptoms included testicular swelling and falling swelling (91.3%), hydrocele testis (31.0%) and testicular pain (23.0%). Ann Arbor stages I-IV accounted for 53.3%, 16.7%, 14.8% and 15.7%, respectively. Diffuse large B-cell lymphoma (DLBCL) cases were higher at 95.5% than NK/T-cell lymphoma cases at 8.2%. Amongst DLBCL cases, 69.3% were non-germinal centre B-cell (GCB) subtype, and 27.6% were GCB subtype. Immunohistochemistry markers showed 95.9% CD3 negative, 94.9% CD10 negative, 94.4% CD20 positive, 88.4% multiple myeloma oncogene-1 (MUM-1) negative, 73.6% B-cell lymphoma-6 (BCL-6) negative and 66.5% BCL-2 positive. Laboratory findings indicated that 70.4% of patients had a tumour proliferating cell nuclear antigen (Ki67) index of ≥80%, 36.0% had increased serum lactate dehydrogenase level and 22.9% had increased serum β2-microglobulin level. PTL is rare, and it often occurs in elderly male patients. Common symptoms include testicular swelling and falling swelling, and the common histological type is DLBCL. Diagnosis should be based on histopathological characteristics and immunohistochemical examination.

Clinical pathological characteristics analysis of ocular adnexal follicular lymphoma

Objective: To investigate the clinical pathological characteristics of ocular adnexal follicular lymphoma (OAFL). Methods: A retrospective case series study was conducted. Clinical data of 10 OAFL patients diagnosed at Tianjin Eye Hospital from January 1990 to May 2022 were collected. The study analyzed general patient information, medical history, site of involvement, imaging, histopathology, and molecular detection. Among them, 7 cases underwent Epstein-Barr virus-encoded small RNA (EBER) and B-cell lymphoma protein 2 (BCL-2)/immunoglobulin heavy chain gene (IgH) translocation gene detection. Treatment and prognosis of patients were followed up. Results: All 10 patients (10 eyes) had unilateral involvement, including 5 males and 5 females, with an age range of 58 (43, 68) years. Clinical manifestations included eyelid swelling, pink conjunctival thickening, painless slow-growing masses in the lacrimal gland area, extraconal muscle cone, conjunctiva, lacrimal sac, or a combination of lacrimal sac and conjunctiva. Among them, 8 cases were primary, and 2 cases were secondary. According to the Ann Arbor staging, 8 cases were stage Ⅰ-Ⅱ E, and 2 cases were stage Ⅲ E. Histopathological grading revealed 6 cases of grade 1-2 and 3 cases of grade 3A. One case showed grade 3B in the lacrimal sac area and grade 1-2 in the conjunctiva. The predominant subtype was follicular in 4 cases, diffuse in 3 cases, and mixed in 2 cases. One case had a mixed subtype involving the lacrimal sac and conjunctiva. All patients expressed positivity for leukocyte differentiation antigens (CD) 20, CD21, and CD23. Nine cases were positive for CD10, with 1 case showing partial CD10 positivity. All patients were positive for B-cell lymphoma protein 6 (BCL-6), and 9 cases were positive for BCL-2. Specific markers CyclinD1 and Multiple Myeloma Oncogene Protein 1 (MUM-1) were negatively expressed in all cases. The Ki-67 proliferation index ranged from 10% to 90%. Molecular detection was performed in 7 patients, with none showing positive EBER in situ hybridization. However, 5 cases exhibited BCL2/IgH gene fusion. Among 7 patients with follow-up data, the median follow-up time was 17 (6, 34) months. Four patients achieved complete remission, 2 had partial remission, and 1 patient died due to lung infection. Conclusions: OAFL is a tumor originating from follicular center B-cells, characterized by positive expression of BCL-2, CD10, and BCL-6. It can involve the lacrimal gland, extraconal muscles, lacrimal sac, and conjunctiva. Patients generally have a favorable prognosis.

Case Report: A very rare case of a Pleural Effusion revealing Multiple Myeloma

Multiple myeloma is a common malignant bone-based disease. Pleural effusions reported in these patients remain rare. It is commonly due to congestive heart disease, pulmonary embolism, nephrotic syndrome or a second neoplasia. The true myelomatous pleural effusion resulting from a direct tumoral invasion of the pleural are extremely rare. We report here the case of a massive pleural effusion revealing multiple myeloma in a 71-year-old patient. The chest ultrasound showed a massive pleural effusion in the left side with a multinodular thickening of the pleura. The medical thoracoscopy showed a grape-cluster appearance. The diagnosis was made by pleural guided biopsy revealing abnormal plasma cells with an intense positive CD 138 (plasma cell marker) and MUM1 (multiple myeloma oncogene1) staining with a light kappa chain in the protein electrophoresis associated with a myeloma. Unfortunately, our patient died one month after the initial diagnosis. We present also a review of the recent literature in order to highlight the clinical presentations of the myelomatous pleural effusion, the diagnostic tools, the therapeutic strategies as well as the outcomes.

Intracranial dissemination in a primary small cell carcinoma of the brain: a case report and literature review.

Primary intracranial small cell carcinoma (SCC) is extremely rare with only 8 previously reported cases. We describe a case of primary intracranial SCC with intracranial metastasis. A 46-year-old man presented with decreased vision and a red and swollen left eye. Brain magnetic resonance imaging (MRI) revealed a heterogeneously enhanced tumor on the left frontal lobe. Preoperative systemic computed tomography (CT), MRI, and positron emission tomography (PET)-CT revealed no extracranial tumors. The tumor on the left frontal lobe was excised. Immunohistochemical staining on the excision showed positivity for CD56, synaptophysin (Syn), cytokeratin (CK), and Ki-67 (30%), and negativity for thyroid transcriptional factor-1 (TTF-1), glial fibrillary acidic protein (GFAP), B-cell lymphoma 6 (Bcl-6), multiple myeloma oncogene 1 (MUM-1), C-Myc, Vimentin, P40, P53, CK7, CD3, CD5, CD20, CD79a, CD10, and CD23. The pathological examination strongly suggested that the tumor was a primary intracranial SCC. One year after the surgery, the patient was readmitted with slurred speech and slow movements. Three well-defined tumors were found in the left upper frontal lobe by brain MRI. Tumor resection was then performed. Further immunohistochemical examination of the excised tissue displayed the same pattern as previously, indicating the recurrence of intracranial SCC in the left frontal lobe. The patient received adjuvant chemotherapy and radiotherapy after the tumor resection. At the 2-year follow-up, he remained asymptomatic.

Case Report: A very rare case of a pleural effusion revealing multiple myeloma

Multiple myeloma is a common malignant bone-based disease. Pleural effusions reported in these patients remain rare. It is commonly due to congestive heart disease, pulmonary embolism, nephrotic syndrome or a second neoplasia. The true myelomatous pleural effusion resulting from a direct tumoral invasion of the pleural are extremely rare. We report here the case of a massive pleural effusion revealing multiple myeloma in a 71-year-old patient. The chest ultrasound showed a massive pleural effusion in the left side with a multinodular thickening of the pleura. The medical thoracoscopy showed a grape-cluster appearance. The diagnosis was made by pleural guided biopsy revealing abnormal plasma cells with an intense positive CD 138 (plasma cell marker) and MUM1 (multiple myeloma oncogene1) staining with a light kappa chain in the protein electrophoresis associated with a myeloma. Unfortunately, our patient died one month after the initial diagnosis. We present also a review of the recent literature in order to highlight the clinical presentations of the myelomatous pleural effusion, the diagnostic tools, the therapeutic strategies as well as the outcomes.

Gastrointestinal: Real-time observation of rectal malignant lymphoma using endocytoscopy for differentiation from adenocarcinoma.

A 25-year-old man was suspected of having advanced rectal cancer by his previous physician and was referred to our hospital for surgery. Colonoscopy was repeated in our hospital before surgery. White-light imaging revealed a semi-circumferential ulcerative lesion in the rectum (Fig. 1a). The lesion was stained with 1% methylene blue and 0.05% crystal violet solution and evaluated using endocytoscopy. Proliferation of swollen, irregular-shaped nuclei with the formation of glandular duct structures was observed (Fig. 1b). Tissue biopsies were taken for flow cytometry and histological assessment. Histopathological examination after hematoxylin and eosin staining revealed infiltration and proliferation of enlarged dysplastic cells, without formation of glandular ducts (Fig. 1c). Immunohistochemistry was performed in response to the flow cytometry results, and was positive for CD20, CD79a, bcl2, and bcl6, but negative for CD3, CD10, multiple myeloma oncogene 1, and Epstein–Barr-virus-encoded small RNA in situ hybridization. Ki-67 index was 90%. The lesion was diagnosed as diffuse large B-cell lymphoma (DLBCL), germinal center B-cell-like type. Malignant lymphomas of the colon account for 0.1%–0.7% of all colorectal malignancies. In the colon, DLBCL is the second most common tumor after mucosa-associated lymphoid tissue lymphoma. DLBCL is often ulcerated and resembles ulcerated colorectal cancer by white-light endoscopic imaging (Fig. 2a). However, differentiation is required for treatment decision-making. Tissue sampling for flow cytometry can be used to quantify expression of proteins associated with prognosis and to detect multidrug resistance, which makes it useful for measuring prognostic indicators in lymphoid neoplasia. Hence, it is important to make a diagnosis of lymphoma on-site to prevent multiple repeat colonoscopies. Endocytoscopy allows real-time, in vivo visualization of atypical cells on the tumor surface and tissue details, including nuclei and glandular lumens, resulting in high diagnostic accuracy for adenocarcinoma or adenomatous lesions. However, there are few reports of the endocytoscopic findings of lymphomas in the gastrointestinal tract. In this case, endocytoscopy (520×, H290ECI; Olympus, Tokyo, Japan) showed a significant difference between DLBCL and adenocarcinoma, with irregular and rough lumens and agglomeration of distorted nuclei strongly stained with methylene blue (Fig. 2b,c).

Precision Medicine for Chronic Endometritis: Computer-Aided Diagnosis Using Deep Learning Model

Chronic endometritis (CE) is a localized mucosal infectious and inflammatory disorder marked by infiltration of CD138(+) endometrial stromal plasmacytes (ESPC). CE is drawing interest in the field of reproductive medicine because of its association with female infertility of unknown etiology, endometriosis, repeated implantation failure, recurrent pregnancy loss, and multiple maternal/newborn complications. The diagnosis of CE has long relied on somewhat painful endometrial biopsy and histopathologic examinations combined with immunohistochemistry for CD138 (IHC-CD138). With IHC-CD138 only, CE may be potentially over-diagnosed by misidentification of endometrial epithelial cells, which constitutively express CD138, as ESPCs. Fluid hysteroscopy is emerging as an alternative, less-invasive diagnostic tool that can visualize the whole uterine cavity in real-time and enables the detection of several unique mucosal findings associated with CE. The biases in the hysteroscopic diagnosis of CE; however, are the inter-observer and intra-observer disagreements on the interpretation of the endoscopic findings. Additionally, due to the variances in the study designs and adopted diagnostic criteria, there exists some dissociation in the histopathologic and hysteroscopic diagnosis of CE among researchers. To address these questions, novel dual immunohistochemistry for CD138 and another plasmacyte marker multiple myeloma oncogene 1 are currently being tested. Furthermore, computer-aided diagnosis using a deep learning model is being developed for more accurate detection of ESPCs. These approaches have the potential to contribute to the reduction in human errors and biases, the improvement of the diagnostic performance of CE, and the establishment of unified diagnostic criteria and standardized clinical guidelines for the disease.

MUM-1 in canine lymphoma: A pilot study.

Expression of interferon regulatory factor 4 (IRF4)/multiple myeloma oncogene-1 (MUM1) in peripheral T-cell lymphoma (PTCL) in people is associated with a poorer survival outcome compared to cases of PTCL lacking MUM1 expression. The aim of this study was to determine whether MUM1 is expressed in canine peripheral T-cell lymphoma not otherwise specified (PTCL-NOS). For comparison, the presence of MUM1 antigen was also investigated in canine diffuse large B cell lymphoma (DLBCL). Nine cases of PTCL-NOS and 9 cases of DLBCL diagnosed by a commercial veterinary diagnostic laboratory were selected. Positive immunohistochemical labeling for MUM1 was observed in PTCL-NOS (2 out of 9 cases) and DLBCL (3 out of 9 cases). These findings suggest that a subset of neoplastic T and B lymphocytes can express MUM1. The role of MUM1 in the biological behavior and outcome of canine lymphoma (CL) requires further investigation on a larger number of cases.

Analysis of pregnancy outcomes in patients with recurrent implantation failure complicated with chronic endometritis.

Chronic endometritis (CE) has been considered to reduce uterine receptivity and negatively affect reproductive outcomes for in vitro fertilization-embryo transfer (IVF-ET) patients, especially for people undergoing recurrent implantation failure (RIF). To investigate the effects of antibiotic and platelet-rich plasma (PRP) therapy on pregnancy outcomes following frozen-thawed embryo transfer (FET) in RIF patients with CE, endometrial specimens of 327 patients with RIF obtained by endometrial scratching during mid-luteal phase were immunostained by multiple myeloma oncogene-1 (Mum-1)/syndecan-1 (CD138). RIF patients with CE were given antibiotics and PRP treatment. According to the Mum-1+/CD138+ plasmacytes after treatment, patients were divided into persistent weak positive CE (+) group, CE (-) group and non-CE group. FET was performed and the basic characteristics and pregnancy outcomes of patients in three groups were compared. Among 327 RIF patients, 117 patients were complicated with CE, the prevalence was 35.78%. The prevalence of strong positive was 27.22% and that of weak positive was 8.56%. 70.94% patients with CE converted to negative after treatment. There was no significant difference in the basic characteristics, including age, BMI, AMH, AFC, infertility years, infertility types, number of previous transplant cycles, endometrial thickness on transplantation day and number of transplanted embryos (p > 0.05); The positive rate of hCG, clinical pregnancy rate and implantation rate in CE (-) group were significantly increased compared with the weak CE (+) group and non-CE group (p < .01), and the live birth rate also improved (p < .05). Whereas, the rate of early abortion in CE (-) group was 12.70%, which was higher than that in weak CE (+) group and non-CE group (p < .05). After multivariate analysis, number of previous failed cycles and CE remained independent factors associated with live birth rate, while only CE remained to be the independent factor of clinical pregnancy rate. It is recommended to perform CE-related examination for patients with RIF. Antibiotic and PRP treatment can significantly improve pregnancy outcomes of patients with CE negative conversion in FET cycle.

Multiple Myeloma with Aberrant CD3 Expression in a Red-Lored Amazon Parrot (Amazona autumnalis).

A 20-year-old, female, red-lored Amazon parrot (Amazona autumnalis) was presented for a 2-week history of weakness. On physical examination, the bird was quiet, fluffed, weak, and had a distended coelom. Radiographic and ultrasound imaging revealed coelomic distention, increased pulmonary parenchymal opacity, renomegaly, dilated intestines, and a thickened ventricular wall. The results of a complete blood cell count indicated the patient was anemic (28%) and had intermediate to large lymphocytes with immature chromatin that were suspected to be neoplastic. Immunocytochemistry on peripheral blood determined that the suspected circulating neoplastic cells were cluster of differentiation (CD) 3+ and occasionally expressed multiple myeloma oncogene 1 (MUM1). Abnormalities from a plasma biochemistry panel were moderate hyperphosphatemia (6.8 mg/dL), marked hyperproteinemia (13.6 g/L), analbuminemia (0 g/dL), and marked hyperglobulinemia (13.6 g/dL). Agarose gel plasma protein electrophoresis documented the presence of albumin (1.2 g/dL) and monoclonal bands which, on reduced lithium dodecyl sulfate polyacrylamide gel electrophoresis, resolved as 60-kd and ∼25-kd bands consistent with immunoglobulin Y heavy and light chains. On the basis of these findings, multiple myeloma was diagnosed. Because of a poor prognosis, the bird was euthanized for postmortem examination. Bone marrow cytology from samples collected during the postmortem examination revealed 17.4% plasma cells and 24% large immature cells with occasional plasmacytoid features. Histopathologic findings included aggregates of neoplastic plasma cells in the bone marrow, spleen, kidney, liver, gastrointestinal tract, muscle, ovary, and brain. The neoplastic cells were strongly immunoreactive for MUM1 and cluster of differentiation 3 (CD3), but negative for CD79a, paired box protein 5, and CD20. This confirmed the clinical diagnosis of multiple myeloma. This report describes an avian immunoglobulin Y-secreting multiple myeloma with aberrant CD3 expression and pseudoanalbuminemia. Aberrant CD3 expression by avian multiple myeloma may explain previously published cases of birds with a monoclonal gammopathy and apparent T-cell lymphoma diagnosed by CD3 immunoreactivity.

Spurious laboratory results associated with immunoglobulin M gammopathy in a dog with multiple myeloma

An 11 year old female-neutered Labrador presented for facial swelling. Clinicopathological abnormalities included hyperglobulinemia, azotemia, hypercalcemia, nonregenerative anemia, thrombocytopenia, and spurious hypoglycemia. Normoglycemia was subsequently confirmed using a cage-side analyzer (AlphaTRAK, Zoetis, UK). Serum and urine protein electrophoresis documented monoclonal (immunoglobulin M) gammopathy with Bence-Jones proteinuria. Computed tomography imaging revealed a monostotic osteolytic bone-lesion, and bone marrow cytology and histopathology documented plasmacytosis with multiple myeloma oncogene 1 / interferon regulatory factor 4 positivity, consistent with multiple myeloma. Infectious disease testing initially indicated seropositivity for Leishmania, Borrelia, and Anaplasma spp.; however, Leishmania PCR (splenic and bone marrow aspirates), and paired serological titers for Borrelia and Anaplasma were negative. Consequently, initial serological results were considered to be false positive because of paraproteinemia-associated assay interference. Chemotherapy (prednisolone and melphalan combination therapy) was initiated, but the dog was euthanased 30 days later because of the development of pericardial effusion. This is a report of spurious serological (and other laboratory) results occurring secondary to monoclonal gammopathy in a dog.

Application of Hans Algorithm for Subcategorization of Diffuse Large B-Cell Lymphoma in Fine-Needle Aspiration Biopsy Cytology

Introduction: Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease with remarkably variable clinical presentation and outcome. Hans algorithm subclassified DLBCL into prognostically distinct molecular subtypes by using immunohistochemistry (IHC). Fine-needle aspiration biopsy cytology (FNABC) is a first-line diagnostic modality in lymphadenopathy. The study aims to perform IHC on FNABC cell blocks (CBs) for subclassifying according to the Hans algorithm and correlate with case-matched histopathology. Methods: This was a retrospective study carried out between January 2017 and December 2019. All DLBCL FNABC cases with CBs and smears and which had follow-up histopathology were included in the study. Detailed cytomorphological evaluation and CD10, B-cell lymphoma 6 (BCL6), and multiple myeloma oncogene 1 IHCs were performed on CBs. The cases are divided into 3 distinct molecular subtypes based on the Hans algorithm as germinal centre B-cell (GCB), activated B-cell (ABC), and unclassified subtypes. The results were compared with the final histopathology. Results: A total of 44 cases were diagnosed as DLBCL, and 33 cases with sufficient material for further IHC were included in the study. Twelve cases were of the GCB type, 19 were of the ABC type, and 2 remained unclassified. Follow-up histopathology was available in 20 cases. Overall, histopathological concordance was found in 95% of cases (19/20). The single discordant case was classified as GCB on FNABC and was ABC on histopathology. Conclusion: FNABC with CBs is an acceptable alternative to biopsy for providing a complete diagnosis of DLBCL as per the current WHO classification.

ALKBH5 Functions As an Oncogene in Multiple Myeloma

Background: RNA N 6-methyladenosine (m 6A) plays a critical role in regulating gene expression and determining cell fate. The dysregulation of m 6A modulators, including α-ketoglutarate-dependent dioxygenase AlkB homolog 5 (ALKBH5), has been reported to promote tumor development through their enzymatic function. However, the functions of mRNA m 6A and its modulators in multiple myeloma (MM) are largely unknown.Methods: We queried publicly available MM datasets to study the expression profile of m 6A modulators (METTL3, METTL14, WTAP, FTO, and ALKBH5) in MM and their relationships with clinical outcomes in patients with MM. Both gain- and loss-of-function studies were performed to investigate the role of ALKBH5 in MM. The cell proliferation assay, colony formation assay, Annexin V apoptosis analysis, and 5-ethynyl-2′-deoxyuridine (EdU) assay were performed to evaluate the functions of ALKBH5 in MM in vitro. Human MM cell line xenograft models were constructed to examine the effects of ALKBH5 knockdown or overexpression on MM growth in vivo. The rescue assay using catalytically inactive mutant ALKBH5-H204A was conducted to determine whether demethylation activity was required for the function of ALKBH5 in MM. Then, we performed RNA sequencing and m 6A sequencing to explore the key targets that mediated ALKBH5 function in MM. We investigated the gene regulatory mechanism of ALKBH5 in MM by m 6A immunoprecipitation assay, RNA immunoprecipitation assay, RNA decay assay, dual-luciferase reporter assay, and so forth. Gene set enrichment analysis and Western blotting were employed to determine the downstream signaling pathways regulated by ALKBH5 and the recognized target.Results: ALKBH5 was overexpressed in MM and associated with a poor prognosis in patients with MM. The increased ALKBH5 expression was required for the survival and growth of MM cells in vitro and in vivo. Mechanistically, m 6A demethylation activity was required for ALKBH5 to exert tumorigenic effects in MM. Tumor necrosis factor (TNF) receptor-associated factor 1 (TRAF1) was identified as a functionally important target of ALKBH5. ALKBH5 regulated TRAF1 expression via affecting mRNA stability of TRAF1 in an m 6A- and YTHDF2-dependent manner. ALKBH5 promoted MM cell growth and survival partly through TRAF1-mediated activation of NF-κB and MAPK signaling pathways.Conclusion: Our findings showed that ALKBH5 played an oncogenic role in MM and highlighted that ALKBH5 could potentially be a novel therapeutic target in MM. DisclosuresNo relevant conflicts of interest to declare.

Follicular Lymphoma Grade 3A Is Different from Grade 1/2 in the Real World

Background: follicular lymphoma (FL) is histologically subdivided into grades 1/2(FL1/2), 3A(FL3A), and 3B(FL3B). FL3B is more like diffuse large B-cell lymphoma (DLBCL), while the clinical approach to FL1/2 and FL3A has been debated. We aim to explore the clinical, biological characteristics and outcomes between them.Methods: A retrospective analysis of 195 de novo FL patients within the same time frame (1999 to 2020) was identified. 141 patients were FL1/2, and 54 patients were FL3A.Results: Comparing with FL1/2, FL3A patients tend to present ECOG≥1, B symptoms, bone marrow involvement, digestive tract involvement, elevated LDH, Ki-67≥30%, CD10 negative, and multiple myeloma oncogene-1(MUM1) positive, while Ann Arbor stage I or II was usually seen in FL1/2. After received CHOP±R (cyclophosphamide, doxorubicin, vincristine and prednisone ±rituximab), the 5-year overall survival (OS) was 89.5% for FL1/2 and 61.1% for FL3A [ HR =3.742(95%CI:1.838-7.620),P <0.0001], the 5-year progression-free survival (PFS) was 62.5 for FL1/2 and 47.6% for FL3A [ HR =2.113(95%CI:1.297-3.443),P =0.003].Conclusion: FL3A is more aggressive both clinically and biologically compared with FL1/2, and more attention should be paid to the difference of treatment between them. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Intestinal Protein Characterisation of SARS-CoV-2 Entry Molecules ACE2 and TMPRSS2 in Inflammatory Bowel Disease (IBD) and Fatal COVID-19 Infection

—The coronavirus SARS-CoV-2 contributes to morbidity and mortality mainly as a result of immune-pathology in the lungs. Recent data has shown multi-system involvement with widespread viral tropism. Here we present a detailed intestinal protein characterisation of SARS-Cov-2 entry molecules ACE2 and TMPRSS2 in patients with inflammatory bowel disease ([IBD]; ulcerative colitis [UC] and Crohn’s disease [CD]) with age- and sex-matched non-IBD controls, and in those with fatal COVID-19 infection. In our dataset, ACE2 and TMPRSS2 displayed a membrane enterocyte staining in the ileum (due to presence of brush border/microvilli) in contrast to a cytoplasmic pattern in the colon. We also showed a high ACE2/low TMPRSS2 expression pattern in the ileum with a reverse trend in the colon. In UC, colonic ACE2 and TMPRSS2 are cytoplasmic in nature, with significantly higher ACE2 staining intensity compared to non-IBD controls. In inflamed and unaffected IBD mucosa, ileal and colonic enterocyte ACE2 and TMPRSS2 expressions are not modified in the histologic presence of inflammation. We observed immune cells within the lamina propria that expressed ACE2 and TMPRSS2, at higher frequencies in IBD when compared to non-IBD controls. These were identified as plasma cells with multiple myeloma oncogene 1/interferon regulatory factor 4 (MUM1/IRF4) expression. We further analysed the gut histology of six fatal COVID-19 cases, with no difference in colonic and ileal ACE2/TMRPSS2 staining (compared to non-IBD controls) and identified ACE2 + lamina propria plasma cells. Of interest, in this COVID-19 cohort, there was no histologic evidence gut inflammation despite known evidence of viral tropism within the enterocytes. Our data provides evidence for tissue expression of entry molecules ACE2 and TMPRSS2 including a close apposition to plasma cells — both pointing towards a role of the gut in the antecedent immune response to SARS-CoV-2 infection.

Spontaneous Disseminated Histiocytic Sarcoma in a Bengal Tiger (Panthera tigris tigris)

A 16-year-old, female, captive Bengal tiger (Panthera tigris tigris) had a history of a recurrent subcutaneous mass. After two attempts at surgical removal, euthanasia was elected during the third surgical attempt due to the degree of neoplastic infiltration. At necropsy, a large subcutaneous mass infiltrated the dermis, subcutis and abdominal wall with metastasis to the lungs, spleen and adrenal glands. Microscopically, the neoplasm formed nodules comprising sheets of round cells with large irregular nuclei. Toluidine blue and Giemsa stains were negative. An immunohistochemistry panel revealed membranous and cytoplasmic labelling with ionized calcium-binding adaptor molecule 1 (IBA1), cytoplasmic labelling with vimentin but no labelling with multiple myeloma oncogene 1 (MUM1), CD20 or pancytokeratin markers. Based on the gross distribution and histological and immunohistochemistry features, a diagnosis of disseminated histiocytic sarcoma was made. To the best of our knowledge, this is the first record of disseminated histiocytic sarcoma in a captive Bengal tiger.

P–337 Mid-Infrared spectroscopy as a real time diagnostic tool for chronic endometritis

Abstract Study question Can we develop a real-time diagnostic tool for chronic endometritis (CE) by using attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy to evaluate biopsies obtained during hysteroscopy? Summary answer A discrimination model based on the absorbance data was developed by machine learning techniques, differentiating between positive and negative CE histopathology with 97% accuracy. What is known already CE is diagnosed in approximately 15% of infertile women who undergo in vitro fertilization (IVF), in 42% of women with recurrent implantation failure (RIF), and in 57.8% of women with RPL. Diagnosis is done by endometrial biopsy, and the presence of plasma cells in the endometrial stroma is the generally accepted histological diagnostic criterion. However, the histological detection of CE is time-consuming and difficult. ATR-FTIR spectroscopy is a non-destructive method that can provide valuable information on biochemical changes that occur during pathological processes, such as inflammation and cancer. Study design, size, duration We performed a prospective study in which fresh biopsies of endometrium were obtained during standard hysteroscopies. Each biopsy was examined by the spectrophotometer and afterward by histopathological analysis in which multiple myeloma oncogene 1 (MUM–1) staining for plasma cells, a marker of CE, was performed. We planned to investigate 80 samples to develop a discrimination model, and another 40 samples for validation of the model. The study was planned to last two years. Participants/materials, setting, methods Women that underwent hysteroscopy as a part of infertility evaluation were recruited. The hysteroscopies and the biopsy evaluation were performed at the same center. A cut-off of 8 MUM–1 positive cells per 10 high power fields (HPF) was set. We compared the spectroscopy analysis of the positive CE group (≥8) and the negative CE group (&amp;lt;8). Machine learning technique was utilized to build discrimination models. Data analysis was performed using Matlab and Unscrambler software packages. Main results and the role of chance We present preliminary results for our study. Forty-two women were recruited from January 2020 until November 2020. Of the 42 measured spectra, three were discarded due to high measurement noise. Of the 39 biopsies, 33 had MUM–1&amp;lt;8 (CE negative group) and 6 had MUM–1≥8 (CE positive group). Measured spectra of tissue smears from CE negative and positive groups differed from each other in the spectral range of 850–990 [cm–1] (p &amp;lt; 0.05). This wavenumber can be associated with the C-H in-plane bend in the alkene group (CnH2n). A discriminant model was developed between the groups using the Principal Component Analysis and Linear Discriminant Analysis techniques. The accuracy obtained by the model was 97%. We divided the 39 hysteroscopies based on the CE signs into 2 groups: “Negative hysteroscopic-CE” and “Positive hysteroscopic-CE”. Positive hysteroscopic signs were micropolyps, strawberry pattern, hyperemia, punctuation, or pale endometrium. Twenty-three samples were taken in the Negative group and 16 samples were taken in the Positive group. However, measured spectra of tissue smears from negative and positive hysteroscopy groups were not significantly different. The correlation coefficient between hysteroscopy groups and MUM–1 score was r = 0.29, meaning that the characteristic signs of CE in hysteroscopy were not correlated to the histopathology. Limitations, reasons for caution First, these are preliminary results and we need to investigate more samples to validate our model. Second, diagnostic criteria for CE are diverse in the literature and we chose 8 MUM–1 positive cells in 10 HPF, a criterion which may not be accepted by all experts in the field. Wider implications of the findings: ATR-FTIR spectroscopy is highly sensitive to molecular changes and has been utilized as a diagnostic tool in a variety of clinical studies. While histopathological results take about two weeks, ATR-FTIR spectroscopy might give us the possibility to diagnose CE in real-time, allowing an immediate initiation of the appropriate treatment. Trial registration number ClinicalTrials.gov Identifier: NCT04197167

Clinicopathological and Immunohistochemical Profile of Mantle Cell Lymphoma: An Institutional Experience.

IntroductionMantle cell lymphoma (MCL) is a biologically aggressive B-cell non-Hodgkin lymphoma (NHL) with distinctive morphologic, immunophenotypic, and molecular characteristics. Differentiation from other chronic lymphoproliferative disorders is essential for prognostication.AimThis paper aims to study the clinicopathological features of MCL with emphasis on immunohistochemical features and disease correlation.MethodTo do so, clinicopathological characteristics from 21 cases of MCL (14 males, seven females, M:F=2:1) diagnosed in the last five years i.e. 2015 to 2020, were retrospectively reviewed and correlated with immunohistochemistry (IHC) data. Particularly those pertaining to cyclin D1, SRY-box transcription factor 11 (SOX11), cluster of differentiation (CD) 5, CD23, MIB E3 ubiquitin protein ligase 1 (MIB1), tumor protein 53 (TP53), c-myelocytomatosis oncogene product (c-MYC), multiple myeloma oncogene 1 (MUM1), mouse double minute 2 homolog (MDM2), and Epstein-Barr virus latent membrane protein 1 (EBV-LMP1) expression with its aberrations.ObservationsThis study shows that MCL constituted 4.2% (21/500) of all NHLs with a mean age of 57.5 years (median 60 years, range 30 to 80 years). The disease was nodal in 19, and extranodal in the remaining two cases. 14 of 21 (67%) had generalized lymphadenopathy and 71% had bone marrow (BM) involvement. The nodal involvement was diffuse in 9/17 (53%), 8/21 (38%) had a blastoid morphology, and an in-situ MCL pattern was not seen in any of the cases selected for the study. Cyclin D1 immunoexpression correlated well with SOX11; CD5-negative in five cases; and CD23-positive in three cases. TP53 and c-MYC expression were noted in 17/19 (89.4%) and 8/17 (47%), respectively. MUM1 registered positive in six cases. None of the cases showed immunopositivity for MDM2 and EBV-LMP1.ConclusionIn essence, this study indicates that morphological and immunophenotypic subclassification of mantle cell lymphoma with a wider panel of IHC markers is essential for understanding disease biology and better prognostication.

Common Genetic Polymorphisms Contribute to the Association between Non-Melanoma Skin Cancer and Chronic Lymphocytic Leukemia

Introduction: Genome-wide association studies (GWAS) have identified thousands of genetic variants associated with numerous traits and diseases. The analysis of shared genetic associations between diseases can help explain previously found epidemiological associations and provide better insight into disease etiology. Epidemiological studies have demonstrated a consistent positive association between chronic lymphocytic leukemia (CLL) and non-melanoma skin cancers (NMSC); however, the reasons for this association are not well understood. We explored the hypothesis that shared genetic risk factors between CLL and NMSC could contribute to the epidemiological association observed between these diseases.Methods: We identified established loci associated with CLL or NMSC with p<5x10-8 based on published results. Single nucleotide polymorphisms (SNPs) were considered independent if they were ≥500 kb apart and had r2<0.05. We examined the association between NMSC loci and CLL risk in a pooled GWAS of CLL including 3,100 cases and 7,667 controls of European ancestry. In parallel, we investigated the association between CLL loci and NMSC risk in a study of 4,242 basal cell carcinoma (BCC) cases and 12,802 controls and also 825 squamous cell carcinoma (SCC) cases and 11,518 controls. Weighted genetic risk scores (wGRS) were constructed separately for each disease. For each SNP, the odds ratios (OR) for the association with NMSC or CLL was extracted from existing literature. In the wGRS, the number of risk alleles was multiplied by the log-OR of that SNP as reported in the literature. We estimated the OR and 95% confidence interval (95% CI) for the associations with individual SNPs and with the wGRS for both CLL and NMSC using logistic regression, adjusting for age, sex, and principal components to account for population substructure.Results: Six of the established genetic loci for CLL and NMSC overlapped such that at least SNPs associated with CLL and NMSC were within a 1 Mb: 2q33.1, 3q28, 5p15, 6p21, 6p25 and 9q21.Of the reported NMSC loci, 4 index SNPs were nominally associated with CLL risk after adjustment for multiple testing (PFDR-adjusted≤0.05): rs12203592 and rs12210050 at 6p25 (IRF4/EXOC2), rs78378222 at 17p13.1 (TP53), and rs191177147 at 3q28 (LPP). The most significant associations were at 6p25.3 region in the IRF4/EXOC2 locus: rs12203592 (OR: 1.41, 95% CI: 1.29-1.54, P=7.34 x 10-14) and rs12210050 (OR=1.23, 95% CI: 1.12-1.35, P=1.97 x 10-5). Using the wGRS from SCC SNPs, we found a pattern of increasing risk of CLL with increasing quartile, reaching statistical significance for quartile 4 vs. quartile 1 (OR=1.26, 95% CI: 1.14-1.39, P=0.001). After excluding the IRF4 polymorphism rs12203592, the association between the wGRS and CLL was no longer significant (OR=1.02, 95% CI: 0.90-1.45, P=0.81).Of the reported CLL SNPs, five were associated with NMSC risk after adjustment for multiple testing (PFDR-adjusted≤0.05): rs9815073 at 3q28 (LPP), rs3769825 at 2q33 (CASP10/CASP8), rs17246404 at 7q31 (POT1), rs4368253 at 18q21 (PMAIP1) and rs872071 at 6p25 (IRF4/EXOC2). The wGRS based on the CLL SNPs was associated with BCC risk (P=0.004) but not with SCC risk (P=0.12), possibly due to a lack of statistical power. Of note, after exclusion of the IRF4 polymorphism, the association between the wGRS and BCC remained statistically significant (P=0.02).Conclusion: We report here a novel and statistically significant association between the genetic burden for risk of NMSC and of CLL. Finally, we uncovered that polymorphisms in the Interferon Regulatory Factor 4 (IRF4) gene appear to contribute to the association between CLL and NMSC. IRF4, also known as multiple myeloma oncogene-1 (MUM1), is a transcriptional regulator expressed in lymphocytes and in melanocytes. Further studies are needed to understand the common biological mechanisms that lead to this association. Overall, these findings highlight the pleiotropic effect of genes in the pathogeneses of these cancers and suggest that they have a partially shared genetic etiology. DisclosuresSmedby:Celgene: Membership on an entity's Board of Directors or advisory committees; Janssen: Research Funding. Shanafelt:Genentech, Celgene, Pharamacyclics, Jansen, GlaxoSmithKline, AbbVie, Hospira, and Cephalon: Research Funding. Cerhan:Janssen: Other: Scientific Advisory Board (REMICADELYM4001); Janssen: Other: Multiple Myeloma Registry Steering.