Exploration of the personalized immune checkpoint atlas of plasma cell dyscrasias patients using high­dimensional single­cell analysis.

Immune checkpoint blockade endows patients with unparalleled success in conquering cancer. Unfortunately, inter­individual heterogeneity causes failure in controlling tumors in many patients. Emerging mass cytometry technology is capable of revealing a multiscale onco­immune landscape that improves the efficacy of cancer immunotherapy. We introduced mass cytometry to determine the personalized immune checkpoint status in bone marrow and peripheral blood samples from 3 patients with multiple myeloma, amyloid light­chain amyloidosis, and solitary bone plasmacytoma and 1 non­hematologic malignancy patient. The expression of 18 immune regulatory receptors and ligands on 17 defined cell populations was simultaneously examined. By single­cell analyses, we identified the T cell clusters that serve as immunosuppressive signal source and revealed integrated immune checkpoint axes of individuals, thereby providing multiple potential immunotherapeutic targets, including programmed cell death protein 1 (PD­1), inducible co­stimulator (ICOS), and cluster of differentiation 28 (CD28), for each patient. Distinguishing the cell populations that function as providers and receivers of the immune checkpoint signals demonstrated a distinct cross­interaction network of immunomodulatory signals in individuals. These in­depth personalized data demonstrate mass cytometry as a powerful innovation to discover the systematical immune status in the primary and peripheral tumor microenvironment.

Activated gp130 signaling selectively targets B cell differentiation to induce mature lymphoma and plasmacytoma.

Aberrant activity of the glycoprotein 130 130/JAK/STAT3 (gp130/JAK/STAT3) signaling axis is a recurrent event in inflammation and cancer. In particular, it is associated with a wide range of hematological malignancies, including multiple myeloma and leukemia. Novel targeted therapies have only been successful for some subtypes of these malignancies, underlining the need for developing robust mouse models to better dissect the role of this pathway in specific tumorigenic processes. Here, we investigated the role of selective gp130/JAK/STAT3 activation by generating a conditional mouse model. This model targeted constitutively active, cell-autonomous gp130 activity to B cells, as well as to the entire hematopoietic system. We found that regardless of the timing of activation in B cells, constitutively active gp130 signaling resulted in the formation specifically of mature B cell lymphomas and plasma cell disorders with full penetrance, only with different latencies, where infiltrating CD138+ cells were a dominant feature in every tumor. Furthermore, constitutively active gp130 signaling in all adult hematopoietic cells also led to the development specifically of largely mature, aggressive B cell cancers, again with a high penetrance of CD138+ tumors. Importantly, gp130 activity abrogated the differentiation block induced by a B cell-targeted Myc transgene and resulted in a complete penetrance of the gp130-associated, CD138+, mature B cell lymphoma phenotype. Thus, gp130 signaling selectively provides a strong growth and differentiation advantage for mature B cells and directs lymphomagenesis specifically toward terminally differentiated B cell cancers.

Improved clonality analysis based on immunoglobulin kappa locus for canine cutaneous plasmacytoma.

Sensitivity of clonality analysis based on immunoglobulin heavy chain (IGH) in canine cutaneous plasmacytoma is lower than that in diffuse large B cell lymphoma (DLBCL) because of somatic hypermutation occurring at the IGH locus. Therefore, this study aimed to improve the sensitivity of clonality analysis for canine cutaneous plasmacytoma. To achieve this, clonality analysis based on the immunoglobulin kappa chain (IGK) locus was established. Sensitivity and specificity were examined in genomic DNA extracted from formalin-fixed paraffin-embedded sections of cutaneous plasmacytomas, DLBCLs, and lymph nodes without lymphoma. Forward primers were designed based on the IGKV genes, and reverse primers were designed based on the IGKJ genes and kappa deleting element (Kde). Analysis using IGKV and IGKJ primers demonstrated clonality in 24 of 29 cutaneous plasmacytomas (82.8%), while analysis with primers for IGKV and Kde showed clonality in 16 of 29 cases (55.2%). In DLBCL, the IGKV and IGKJ primer set yielded clonality in 18 of 23 cases (78.3%), and the IGKV and Kde primer set yielded 9 of 23 cases (39.1%). No clonal results were obtained from 23 lymph nodes without lymphoma. Sensitivity of the IGKV and IGKJ primer set was significantly higher than that of the IGH primers reported previously. Thus, clonality analysis based on the IGK locus can be utilized for canine B cell tumors. In conclusion, clonality testing based on IGH and IGK may be beneficial as an adjunct tool for diagnosis of canine B cell tumors including cutaneous plasmacytoma.

Reactivity with the EpCAM-specific antibodies MOC-31 and Ber-Ep4 in plasma cell neoplasms: a potential diagnostic pitfall in cytology samples.

OBJECTIVE: Epithelial cell adhesion molecule (EpCAM) is a protein expressed on surfaces of healthy epithelia, and is overexpressed in dysplasias and carcinomas. Immunohistochemistry (IHC) utilizing antibodies that react with EpCAM, such as MOC-31 and Ber-EP4, distinguish reactive mesothelial cells from carcinomas in serous effusions. IHC is crucial in effusions with singly dispersed atypical cells, a scenario with a broad differential, including hematopoietic malignancies. Plasma cell neoplasms (PCN) are the second most common hematopoietic malignancy, manifesting as multiple myeloma or plasmacytoma, with 6% of cases developing serous cavity involvement. Most PCNs are readily recognizable; however, variants that deviate from the classic cytomorphology risk erroneous diagnosis. This study demonstrates EpCAM expression in a subset of PCNs, highlighting a potential diagnostic pitfall in serous effusion cytology. METHODS: A 10-year retrospective search for cytology specimens with a diagnosis of PCN was performed. All cases demonstrating CD138/CD38 and monoclonal immunoglobulin expression, and adequately cellular cell block were included. IHC analysis for MOC-31 and Ber-EP4 was performed using Ventana Benchmark Ultra. Scoring was performed as follows: total IHC score equals the positive proportion (0 = no positive tumor cells; 1 = <1%; 2 = 1-10%; 3 =11-33%; 4 = 34-66%; 5 = 67-100%) plus staining intensity (0, no staining; 1, weak; 2, moderate; 3, strong). A score > 4 was considered positive. RESULTS: 2 of 28 (7%) PCNs demonstrated positivity for MOC-31 and Ber-Ep4. CONCLUSION: A subset of PCNs in cytology samples show positivity for MOC-31 and Ber-EP4 which could result in misinterpretation as carcinoma.

Primary plasmablastic plasmacytoma in the stomach of an immunocompetent adult: A case report.

RATIONALE: Extramedullary plasmacytomas (EMP) are tumors composed by a monoclonal population of plasma cells that arise in extraosseus tissues, occupying <5% of all plasma cell neoplasms. Gastrointestinal solitary extramedullary plasmacytoma is rare, just comprises about 5% of all EMPs. The most common site is small intestine. The gastric incidence is much rare and especially the plasmablastic plasmacytoma in the stomach. PATIENT CONCERNS: A 65-year-old man had an epigastric discomfort and mass for about 2 months. Gastroscopy revealed a malignant tumor in the gastric body. Abdomen computed tomography (CT) showed that the gastric cavity was filled, and the irregular soft tissue shadow was seen in the greater curvature, and the enhancement was obvious. To get more tissue, we conducted stomach puncture biopsy. Pathology showed small-round cell malignant tumors. And immunohistochemical examinations revealed that the tumor tend to be a plasma cell tumor. DIAGNOSIS: Gastric plasma cell tumor. INTERVENTIONS: Distal gastrectomy was performed to treat the tumor. OUTCOMES: In addition to ascites caused by hypoproteinemia, there were no postoperative complications. Postoperative pathologic report showed plasmablastic plasmacytoma. Histopathologic examination of the specimen revealed plasmablastic plasmacytoma originating in the stomach with transmural extension, but without lymph node metastasis. The patient is regularly followed up at a postoperative clinic and is doing well, and at present there is no plan for adjuvant treatment. LESSONS: Surgical resection is good option for gastrointestinal EMP.

[A case report of cutaneous plasmacytosis].

A collection of plasma cells in the skin can represent a broad spectrum of disease entities. Secondary syphilis, primary cutaneous plasmacytoma, primary cutaneous plasmacytosis, cutaneous lymphoid hyperplasia and nodular amyloidosis are considered possible differential diagnoses. The primary cutaneous plasma cell disorders can range from malignant to benign plasma cell neoplasms. The malignant conditions are neoplastic diseases having monoclonal proliferations, rapid progression and fatal outcome while the benign plasma cell disorders usually show polyclonality, chronicity and benign process, including plasmacytosis. We present a case of cutaneous plasmacytosis. The patient was a 34-year-old man, presented with disseminated reddish-brown plaques and nodules on the right side of the hips, inguinal groove, and the thigh. Histopathologically, mature plasma cells perivascular infiltrates were observed mainly in the dermis. Polyclonality of infiltrating plasma cells with coexistence of both kappa and gamma chain-positive cells demonstrated with immunohistochemistry, as well as CD20+++, CD38++++, CD79a++++, CD138++, Ki67<30%. The diagnosis, cutaneous plasmacytosis, was established by the pertinent laboratory findings. Primary cutaneous plasmacytosis was an uncommon reactive lymphoplasmacytic disorder of uncertain etiology. Cutaneous plasmacytosis is a rare disease characterized by peculiar multiple eruptions and hyper gamma globulinemia. It has been mainly described in patients of Japanese descent, with only few reports in Caucasians and Chinese, although information concerning the disorder was limited to individual case reports. Cutaneous plasmacytosis is a rare disorder, which is characterized by multiple red to dark-brown nodules and plaques on the trunk and usually associated with polyclonal hyper gamma globulinaemia. Primary cutaneous plasmacytosis or cutaneous plasmacytosis was thought to be a reactive process with unknown etiology. Histologically, lesions contain dense perivascular infiltration of mature polyclonal plasma cells without any atypia, in the dermis and subcutaneous fat. The clinical course is chronic and benign without spontaneous remission. Available treatments for cutaneous plasmacytosis include psoralen ultraviolet A radiotherapy, systemic chemotherapy and intralesional steroid injection. The patient with cutaneous plasmacytosis in this report was treated with tacrolimus ointment and psoralen ultraviolet A.

Kaposi's Sarcoma-Associated Herpesvirus Latency Locus Renders B Cells Hyperresponsive to Secondary Infections.

Kaposi's sarcoma-associated herpesvirus (KSHV) induces B cell hyperplasia and neoplasia, such as multicentric Castleman's disease (MCD) and primary effusion lymphoma (PEL). To explore KSHV-induced B cell reprogramming in vivo, we expressed the KSHV latency locus, inclusive of all viral microRNAs (miRNAs), in B cells of transgenic mice in the absence of the inhibitory FcγRIIB receptor. The BALB/c strain was chosen as this is the preferred model to study B cell differentiation. The mice developed hyperglobulinemia, plasmacytosis, and B lymphoid hyperplasia. This phenotype was ameliorated by everolimus, which is a rapamycin derivative used for the treatment of mantle cell lymphoma. KSHV latency mice exhibited hyperresponsiveness to the T-dependent (TD) antigen mimic anti-CD40 and increased incidence of pristane-induced inflammation. Lastly, the adaptive immunity against a secondary infection with Zika virus (ZIKV) was markedly enhanced. These phenotypes are consistent with KSHV lowering the activation threshold of latently infected B cells, which may be beneficial in areas of endemicity, where KSHV is acquired in childhood and infections are common.IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) establishes latency in B cells and is stringently linked to primary effusion lymphoma (PEL) and the premalignant B cell hyperplasia multicentric Castleman's disease (MCD). To investigate potential genetic background effects, we expressed the KSHV miRNAs in BALB/c transgenic mice. BALB/c mice are the preferred strain for B cell hybridoma development because of their propensity to develop predictable B cell responses to antigen. The BALB/c latency mice exhibited a higher incidence of B cell hyperplasia as well as sustained hyperglobulinemia. The development of neutralizing antibodies against ZIKV was augmented in BALB/c latency mice. Hyperglobulinemia was dampened by everolimus, a derivative of rapamycin, suggesting a role for mTOR inhibitors in managing immune activation, which is hallmark of KSHV infection as well as HIV infection.

[Tumors of unknown origin: Case report].

IgG4 related disease is a recently recognized chronic fibrotic, inflammatory condition, caused by infiltrating IgG4 positive plasma cells that can cause tumor like disease in almost any organ in the body. Typical histopathology is lymphoplasmocytic infiltration of IgG4 positive cells, storiform fibrosis and obliterative phlebitis. Glucocorticoids alone or in combination with B-cell depletion with rituximab causes often good, lasting response. We present here a lady with recurrent lung infiltration that simulated pneumonia and later tumor of the lung. She was also earlier diagnosed with lump in the breast that was found to contain similar IgG4 positive plasma cells that was also demonstrated in the lung biopsy. She responded very well to rituximab given on 2 occasions. Three years after this treatment she is in total remission. Key words: IgG4 related disease, rituximab treatment, plasmacytoma of breast, tumor of lung Correspondence: Arni Jon Geirsson, arnijon@landspitali.is.

Oncolytic Virotherapy: A Contest between Apples and Oranges.

Viruses can be engineered or adapted for selective propagation in neoplastic tissues and further modified for therapeutic transgene expression to enhance their antitumor potency and druggability. Oncolytic viruses (OVs) can be administered locally or intravenously and spread to a variable degree at sites of tumor growth. OV-infected tumor cells die in situ, releasing viral and tumor antigens that are phagocytosed by macrophages, transported to regional lymph nodes, and presented to antigen-reactive T cells, which proliferate before dispersing to kill uninfected tumor cells at distant sites. Several OVs are showing clinical promise, and one of them, talimogene laherparepvec (T-VEC), was recently granted marketing approval for intratumoral therapy of nonresectable metastatic melanoma. T-VEC also appears to substantially enhance clinical responsiveness to checkpoint inhibitor antibody therapy. Here, we examine the T-VEC paradigm and review some of the approaches currently being pursued to develop the next generation of OVs for both local and systemic administration, as well as for use in combination with other immunomodulatory agents.

Morphology and immunoreactivity of canine and feline extramedullary plasmacytomas.

The aim of the study was the evaluation of morphology and immunophenotype of canine (19 cases) and feline (7 cases) extramedullary plasmacytomas. Tumours, located in skin, oral cavity and spleen were surgically excised, fixed and processed for histopathology and immunohistochemistry (CD79α, CD18, proliferating cell nuclear antigen, metallothionein). Histologically, tumours were classified into mature, cleaved, asynchronous, polymorphous blastic, hyalin, or monomorphous blastic type. All evaluated tumours showed cytoplasmic expression of CD79α antigen. The expression of CD18 was observed in canine cutaneous and splenic tumours. In canine tumours expression of metallothionein was low to moderate, while in feline plasmacytomas - absent or low. In canine tumours, the mitotic index and proliferating cell nuclear antigen index were positively correlated with the expression of metallothionein. In feline tumours no correlation between mitotic index, proliferating cell nuclear antigen and metallothionein was found. This is the first study describing expression of metallothionein in canine and feline extramedullary plasmacytoma.

Serum-free light-chain analysis in diagnosis and management of multiple myeloma and related conditions.

The introduction of the serum-free light-chain (S-FLC) assay has been a breakthrough in the diagnosis and management of plasma cell dyscrasias, particularly monoclonal light-chain diseases. The first method, proposed in 2001, quantifies serum-free light-chains using polyclonal antibodies. More recently, assays based on monoclonal antibodies have entered into clinical practice. S-FLC measurement plays a central role in the screening for multiple myeloma and related conditions, in association with electrophoretic techniques. Analysis of S-FLC is essential in assessing the risk of progression of precursor diseases to overt plasma cell dyscrasias. It is also useful for risk stratification in solitary plasmacytoma and AL amyloidosis. The S-FLC measurement is part of the new diagnostic criteria for multiple myeloma, and provides a marker to follow changes in clonal substructure over time. Finally, the evaluation of S-FLC is fundamental for assessing the response to treatment in monoclonal light chain diseases.

Multiple myeloma and multiple plasmacytomas associated with free gamma heavy chain, free kappa light chain and IgGk paraproteins: an unusual triple gammopathy.

Multiple myeloma is a malignant plasma cell dyscrasia that is becoming more prevalent in an increasingly ageing population. It is a complex disease with clinical phases ranging from the premalignant monoclonal gammopathy of undetermined significance to asymptomatic (smouldering) myeloma and then symptomatic myeloma; the latter occasionally terminating in the clonal proliferation of plasma cells outside the bone marrow. We present a patient whose clonally evolved disease from monoclonal gammopathy of undetermined significance to multiple myeloma demonstrated the presence of an unusual combination of monoclonal immunoproteins. Capillary electrophoresis demonstrated the presence of three paraproteins in the gamma region (γ-region), two of which were additional to the IgGk paraprotein which migrated in the slow γ-region at initial diagnosis. Subsequent isotypic identification of the new paraproteins was not possible by immunotyping and initial immunofixation studies failed to definitively characterize the monoclonal proteins. After reduction with beta-mercaptoethanol, two paraproteins were detected by both capillary and gel electrophoresis. However, only immunofixation was able to resolve three distinct monoclonal bands, confirming the presence of free monoclonal kappa light chains in the mid-gamma region and free monoclonal heavy chains in the fast gamma region. Triple gammopathies in themselves are uncommon; this case presents a very unusual combination of paraproteins which required various electrophoretical and immunochemical techniques to identify and characterize them. The change of electrophoretic signature from the monoclonal gammopathy of undetermined significance phase to the diagnosis of multiple myeloma suggested that a number of genetically distinct subclones were present in the pretreatment clonal evolution of the disease.

B-cell Non-Hodgkin Lymphomas with Plasmacytic Differentiation.

B-cell non-Hodgkin lymphomas with plasmacytic differentiation are a diverse group of entities with extremely variable morphologic features. Diagnostic challenges can arise in differentiating lymphoplasmacytic lymphoma from marginal zone lymphoma and other low-grade B-cell lymphomas. In addition, plasmablastic lymphomas can be difficult to distinguish from diffuse large B-cell lymphoma or other high-grade lymphomas. Judicious use of immunohistochemical studies and molecular testing can assist in appropriate classification.

Plasmacytoma-like Posttransplant Lymphoproliferative Disorder in a Pediatric Heart Transplant Recipient.

Posttransplant lymphoproliferative disorder (PTLD) is a diversely manifesting group of lymphoid or plasmacytic proliferations found in solid organ and bone marrow transplant recipients. PTLD occurs as a result of immunosuppression and is often driven by the Epstein Barr virus. Although most commonly of B-cell origin, similar to B-cell lymphomas, PTLD can rarely present as a plasmacytic process, resembling multiple myeloma. Although more common in adults, 8 cases of plasmacytoma-like PTLD have been reported in pediatric renal and combined small bowel-liver transplant recipients. Here, we present a rare report of a plasmacytoma-like PTLD case in a pediatric heart transplant recipient.

Utility of CD54, CD229, and CD319 for the identification of plasma cells in patients with clonal plasma cell diseases.

BACKGROUND: Multiparameter flow cytometry (MFC) identification and characterization of plasma cells (PCs) is a useful tool to support diagnosis, prognostication, and monitoring of PC diseases (PCD). Currently, the number of MFC markers suited for the identification of PC remains limited. Moreover, antibody therapies against PC-associated markers further compromise the utility of the most widely used reagents (e.g., CD38). Despite markers other than CD38 and CD138 are recognized as potentially useful PC-identification markers, no study has comparatively evaluated their performance in combination with CD38 and CD138. Here we compared the utility of CD229, CD54, and CD319 for the identification of normal and aberrant PCs. METHODS: Bone marrow (BM) samples from 5 healthy controls, two noninfiltrated nonHodgkin lymphoma cases and 46 PCD patients plus 3 extraosseous plasmocytomas, and normal peripheral blood (PB) specimens, were studied. RESULTS: Our results showed adequate performance of all three markers once combined with CD38. In contrast, when combined with CD138 for the identification of PC, only CD229 provided a good discrimination between PCs and all other cells for all BM and PB samples analyzed; in contrast, CD54 and CD319 showed limited utility for the identification of PCs, mainly because of significant overlap of the staining for these two markers on PCs and other myeloid cells in the sample. CONCLUSIONS: From the three markers evaluated, CD229 may be considered as the most reliable marker to replace CD38 or CD138 for the identification of PCs in patients undergoing anti-CD38 or anti-CD138 therapy, until a better alternative is available.