Mature Plasma Cells Research Articles

FAM46C controls antibody production by the polyadenylation of immunoglobulin mRNAs and inhibits cell migration in multiple myeloma.

FAM46C, frequently mutated in multiple myeloma (MM), has recently been shown to encode a non‐canonical poly(A) polymerase (ncPAP). However, its target mRNAs and its role in MM pathogenesis remain mostly unknown. Using CRISPR‐Cas9 technology and gene expression analysis, we found that the inactivation of FAM46C in MM down‐regulates immunoglobulins (Igs) and several mRNAs encoding ER‐resident proteins, including some involved in unfolded protein response and others that affect glycosylation. Interestingly, we show that FAM46C expression is induced during plasma cell (PC) differentiation and that Ig mRNAs encoding heavy and light chains are substrates of the ncPAP, as revealed by poly(A) tail‐length determination assays. The absence of the ncPAP results in Ig mRNA poly(A) tail‐shortening, leading to a reduction in mRNA and protein abundance. On the other hand, loss of FAM46C up‐regulates metastasis‐associated lncRNA MALAT1 and results in a sharp increase in the migration ability. This phenotype depends mainly on the activation of PI3K/Rac1 signalling, which might have significant therapeutic implications. In conclusion, our results identify Ig mRNAs as targets of FAM46C, reveal an important function of this protein during PC maturation to increase antibody production and suggest that its role as a tumour suppressor might be related to the inhibition of myeloma cell migration.

Morphological Study of Mesenteric Lymph Nodes in Rats with Experimental Breast Cancer Treated with Fragmented Double-Stranded DNA.

We performed a morphometric analysis of mesenteric lymph nodes in rats with breast cancer induced by administration of N-methyl-N-nitrosourea. The volume of the paracortical zone and the number of mature plasma cells in the medullary sinuses were increased and the volume of lymphoid nodules with germinal centers and the number of macrophages were decreased in the group with tumor resection and chemotherapy in comparison with untreated rats with breast cancer. In rats receiving fragmented human double-stranded DNA in combination with adjuvant therapy, the volume of marginal and medullary sinuses and the number of small lymphocytes and macrophages in the paracortical zone increased in comparison with the group receiving chemotherapy without DNA preparation; the volume of lymphoid nodules with germinal centers returned to the level observed in the intact group; the volume of medullary substance and proliferative activity of cells in the germinal centers and medullary substance decreased, the number of mature plasma cells returned to normal in the medullary substance and decreased in the medullary sinuses.

Genetic Basis of Extramedullary Plasmablastic Transformation of Multiple Myeloma.

In patients with multiple myeloma, plasmablastic transformation in the bone marrow is rare and associated with poor outcomes. The significance of discordant extramedullary plasmablastic transformation in patients with small, mature clonal plasma cells in the bone marrow has not been well studied. Here, we report the clinicopathologic, cytogenetic, and molecular features of 10 such patients (male/female: 6/4, median age: 65 y, range: 48 to 76 y) with an established diagnosis of multiple myeloma in the bone marrow composed of small, mature plasma cells in parallel with a concurrent or subsequent extramedullary plasmablastic transformation. Eight patients with available survival data showed an overall aggressive clinical course with a median survival of 4.5 months after the diagnosis of extramedullary plasmablastic transformation, despite aggressive treatment and even in patients with low-level bone marrow involvement. Pathologically, the extramedullary plasmablastic myeloma were clonally related to the corresponding bone marrow plasma cells, showed high levels of CMYC and/or P53 expression with a high Ki-67 proliferation index by immunohistochemistry and harbored more complex genomic aberrations including frequent mutations in the RAS pathway and MYC rearrangements compared with their bone marrow counterparts. In summary, although genetic and immunohistochemical studies were not uniformly performed on all cases due to the retrospective nature of this study, our data suggest that discordant extramedullary plasmablastic transformation of multiple myeloma has an aggressive clinical course and is characterized by frequent mutations in the RAS pathway and more complex genomic abnormalities.

Plasma Cell-Rich Acute Rejection in Renal Transplantation: A Case Report

Plasma cell-rich acute rejection (PCAR) is a rare type of allograft rejection in renal transplantation. It is characterized by the presence of mature plasma cells that compromise more than 10% of inflammatory cells infiltrating the renal graft. The pathogenesis of PCAR is unknown, appears late, and has been related mainly to insufficient immunosuppression or infections. The treatment is not clearly defined, and the graft survival is poor.Here, we report a case series of 3 Spanish patients diagnosed with PCAR accompanied by donor-specific antibodies (DSA) after kidney transplantation. Mean to diagnosis was 2-12 years post-transplantation, and they began with abrupt deterioration of renal function. All patients were women and had preceding viral infection. In addition, two of the three patients recognize a doubtful adherence to immunosuppression.About treatment, 2 of the 3 patients, because the biopsy of the renal graft showed signs suggestive of incipient antibody-mediated rejection (ABMR) (glomerulitis, capilaritis, transplant glomerulopathy), were started with corticosteroids, anti-thymoglobulin, plasmapheresis, and intravenous immunoglobulins. The last patient, who only showed PCAR at biopsy, was treated with corticosteroids and anti-thymoglobulin.After treatment, graft function improved in all of them, but one patient developed an ABMR and another required a dialysis program, all of which indicates the difficulty in management and treatment of PCAR.

Correlation Analysis of Morphological Changes of Structural and Functional Areas of Mesenteric Lymph Nodes with Cytokines of Lymph of Thoracic Lymphatic Duct in Experimental Breast Cancer.

We performed a correlation analysis of the morphometric parameters of mesenteric lymph nodes and cytokine content in the lymph of thoracic duct in rats with chemically induced breast cancer. The study showed that activity of the local immune response in the lymph nodes in breast cancer is aimed at antitumor protection. In breast cancer, the area of the paracortical zone remained at the level of the intact group, while the area of lymphoid nodules with germinative centers and the area of medullary substance increased; the number of macrophages in the thymus-dependent zone and zone responsible for humoral immunity also increased. The following positive correlations were revealed: in germinative centers and medullary substance, number of mitotic cells correlated with cytokine IL-5 content and the number of medium lymphocytes correlated with the content of chemokine MIP-1α; in the germinative centers, the number of immunoblasts correlated with the level of cytokine GRO/KC, in the paracortical zone, the number of macrophages correlated with the level of chemokine MCP-1, the number of reticular cells correlated with IL-6 and M-CSF content; in medullary substance, the number of small lymphocytes and mature cells plasma cells (their content was reduced) correlated with the level of chemokine GRO/KC, which can be caused by their migration from the lymph node.

Tumor miofibroblástico inflamatorio de la mama: presentación de caso y revisión de la literatura

Inflammatory myofibroblastic tumour (IMT) belongs to the group of pseudo inflammatory tumours, composed of fusiform cells mixed with mature plasma cells and inflammatory cells. It is frequently described in the abdomen, pelvis, lung and retroperitoneum in paediatric and adolescent patients. Presentation in the mammary gland is extremely rare (27 cases described in the literature), which is why its natural history, recurrence and metastatic potential remain under study. We present the case of a postmenopausal patient, who was treated with conservation surgery, as well as a review of the literature of this entity.

Combined membranous glomerulonephritis and plasma cell-rich acute rejection in renal transplant recipient presented as nephrotic syndrome: case report and review of literature

Despite the contentious trials to improve the rates of acute rejection episodes and improve the renal allograft survival with potent immunosuppressants, the occurrence of more than 10 % of the inflammatory cells infiltrating renal allograft as mature plasma cells is uncommon and was recognized as plasma cell rich acute rejection (PCAR). Combination of PCAR with other glomerulopathies has not been reported before. Here, we report a case of late-onset PCAR associated with membranous glomerulonephritis (MGN) diagnosed at an early stage (2 years after transplantation). A 58-year-old male patient had end-stage kidney disease secondary to diabetic nephropathy and he was maintained on hemodialysis for 2 years until he underwent overseas living kidney transplantation. He presented to our center in Kuwait on the sixth day postoperatively, with stable renal function. He was maintained on steroid, cyclosporine, and mycophenolate mofetil. Two years after transplant, he developed picture of nephritic syndrome. His graft biopsy showed plasma cell-rich interstitial infiltrate associated with mild edema and focal tubulitis (PCACR) in addition to MGN. Further immunohistochemistry tests revealed both B-cell and T-cell markers (CD20 and CD3) were expressed in the lymphoplasmacytic infiltrate with predominant T lymphocytes. The possibility of multiple myeloma was ruled out. He received pulse steroid 1 g od for 3 days. His maintenance immunosuppression was intensified to tacrolimus-based regimen. Cluster differentiated lymphocyte count showed high CD-19 cells; therefore, we gave him a single dose of rituximab (375 mg/m2). Follow-up graft biopsy (1 month later) revealed MGN with complete resolution of plasma cells. His proteinuria started to improve after 4 months of management. He is enjoying stable graft function with controlled diabetes mellitus. PCAR is a treatable form of acute cellular rejection, and its combination with MGN will need special care with specific CD20 ablation therapy.

Mediastinal plasmocytoma with multiple myeloma: unusual presentation

We report a rare case about a 60 years old Moroccan woman with a past medical history of gastrointestinal tuberculosis, admitted to the emergency department with fever, coughing, weight loss, Spontaneous sternum fracture, back and chest pain. A physical examination revealed diffuse bone pain on palpation. Laboratory testing revealed anemia at 9 g/dl, high serum viscosity (80 mm/h). C reactive protein at 100 mg/l. Serum protein electrophoresis showed monoclonal spike (30 g/dL) in the gamma zone. Serum immunofixation study revealed a IgG lambda monoclonal paraprotein. Serum free light chains (kappa/lambda ratio: 0,04). Total protein level (88g/L). 24-h proteinuria was negative, complete metabolic panel, blood calcium and serum creatinine were normal. Bone marrow aspiration was performed and showed mature and dystrophic plasma cells at 38 %. CT scan of the chest revealed left broncho pulmonary focus associated to a sternal fracture (A), mediastinal tissue mass of superior and posterior mediastinum of 30mm diameter (B), scanner-guided biopsy confirmed plasmocytoma, bronchoscopy with biopsy which objectified an inflammatory lesion. Hematological assessment concluded to mediastinal plasmocytoma with multiple myeloma Ig G Lambda. The Revised International Staging System is at 2. The patient received empirical antimicrobial therapy with good response, then she was treated with VTD protocol (Bortezomib, thalidomide and dexamethasone), achieving complete remission followed by autologous stem cell transplantation (C,D). Consolidation therapy and maintenance are planned to maintain remission.

A Maturation Index Defines Newly Diagnosed Multiple Myeloma Patients with Advanced Immunophenotypic and Molecular Differentiation Profiles Associated with Poor Prognosis

A Maturation Index Defines Newly Diagnosed Multiple Myeloma Patients with Advanced Immunophenotypic and Molecular Differentiation Profiles Associated with Poor Prognosis

First Clinical Study of the B-Cell Maturation Antigen (BCMA) 2+1 T Cell Engager (TCE) CC-93269 in Patients (Pts) with Relapsed/Refractory Multiple Myeloma (RRMM): Interim Results of a Phase 1 Multicenter Trial

First Clinical Study of the B-Cell Maturation Antigen (BCMA) 2+1 T Cell Engager (TCE) CC-93269 in Patients (Pts) with Relapsed/Refractory Multiple Myeloma (RRMM): Interim Results of a Phase 1 Multicenter Trial

Metastatic Lymphoepithelioma-like Carcinoma Treated with PD1-Antibody:Is Epstein-Barr Virus Eradication the Main Mechanism in 2 Refractory Cases in Children?

Background: Lymphoepithelioma-like carcinoma (LELC) is a rare high-grade carcinoma that can occur throughout the body. It is usually associated with Epstein-Barr virus (EBV) infection in patients from Asian countries. Pulmonary LELC occurs mostly in Asian women,, in their fifth or sixth decade, with no previous history of smoking. Optimal treatment has not been clearly established. Treatment options are based on surgery for early stage and on multimodal therapy for advanced stages.There is no report on children treated with programmed cell death 1 (PD1) inhibitor. Case Description: Case 1, a 12-year-old female who was admitted for cough and serious neck swelling. PET-CT showed multiple lymph nodes enlargement in the whole body ;among them, the anterior mediastinal lesions were fused with each other and the boundary between adjacent large vessels was unclear; multiple lung and pleural metastasis . EB virus in blood was 7.48×104copies/mL.Bone marrow morphology: Naive lymphocytes account for 2.5%, some lymphocytes are irregular; Mature plasma cells easy to see.Bone marrow immunotypes: 0.56% CD19-CD5-CD10-abnormally mature B lymphocytes. Biopsy: (left neck lump) LELC, EBERs(+).Partial remission was achieved after 2 courses of paclitaxel / carboplatin/ apatinib protocal. Due to delay with varicella, the tumor came back. After the third course of treatment,she showed chest tightness and pleural effusion, EB virus in blood increased, PDL1 protein was TPS 80% positive. After one PD1 antibody combined with chemotherapy,lymph nodes and thoracic lesions significantly reduced (Image1-3), blood EB virus turned to negative. She is now continuing with PD1 inhibitor. Case 2 , 10-year-old boy with huge right mouth bottom and right neck mass was diagnosed as LELC with EBER(+), TP53 (3%, +), CMYC (80% +). After chemotherapy, surgery and radiotherapy,he achieved complete remission. However, multiple metastases in mediastinum lymph nodes, pleura and lung happened and surgery, chemotherapy followed by automatic stem cell transplantation and pulmonary radiotherapy were given. CT-led puncture biopsy of suspected pulmonary residual lesion showed chronic inflammation with the mild growth of the alveoli, no malignant tumor.The child took oral Tegafur as maintenance regimen. 2 months later, recurrence appeared in lymph nodes, lung ,pleural and acetabular with positive EB virus in pleural effusion. Gene detection with NGS, which includes 312 gene-wide exon region, 208 gene hotspots and 16 fusion genes, did not show mutations such as gene mutations, amplification, and fusion with a clear clinical correlation. Tumor mutation load (TMB) was low, however,the boy had attained good response after PD1 antibody followed by rescue regimen and negative blood EB virus .The 2 patients didn't develop side effects correlating with PD1 inhibitor. Conclusion: To our knowledge, this is the first report of the use of PD1 inhibitor in children with metastatic LELC. These 2 refractory cases were all Epstein-Barr virus (EBV)-driven LELC,are more likely to respond to PD-1 blockade regardless of PD1 expresssion or TMB. EB virus burden disappeared after PD1 blockade,which perhaps can explain the anti-tumor effect.More cases are needed to verify the potential benefits of PD1 inhibitors to treat refractory LELC in children. Figure Disclosures No relevant conflicts of interest to declare.

IRF4 Activity Is Required in Established Plasma Cells to Regulate Gene Transcription and Mitochondrial Homeostasis

The transcription factor interferon regulatory factor 4 (IRF4) is critical for the development, maintenance, and function of plasma cells. The mechanism by which IRF4 exerts its action in mature plasma cells has been elusive due to the death of all such cells upon IRF4 loss. While we identify apoptosis as a critical pathway for the death of plasma cells caused by IRF4 loss, we also determine that IRF4 did not regulate the intrinsic apoptotic pathway directly. By using an inducible IRF4 deletion system in the presence of the overexpression of anti-apoptotic BCL2, we identify genes whose expression is coordinated by IRF4 and that in turn specify plasma cell identity and mitochondrial homeostasis.

Age Related Changes in the Cellular Population and Lymphoid Tissue in the Harderian Gland of Turkey

The aim of the current investigation was to identify the cellular changes of the cell populations positioned within the connective tissue of the harderian glands in Turkeys at different ages extended from one day hatched offspring till the age of seven months. To conduct such research, 50 specimens of harderian glands from the above-dated ages of turkey were collected. Post euthanasia and dissection of the birds, specimens were cut and fixed in 10% neutral buffered formalin and Bouin’s solution. The subsequent routine histological technique was made on these specimens such as dehydration, clearing, paraffin embedding, block preparation, and sectioning. Tissue sections were obtained and stained with different stains such as hematoxylin-eosin, Masson’s trichrome, periodic acid-Schiff (PAS) and Alcian blue (AB). Microscopic findings showed connective tissue of the harderian gland was populated by three types of cells that were heterophils, lymphocytes and plasma cells. The incidence of each of these cells was changed with advance of the age. During the growth of the birds, a gradual shift was taking place in the domination of these three cell types. The lymphoid nodules were detected only in 40 days old poults, whilst in adult birds the harderian glands were contained the largest number of mature plasma cells. Some plasma cells enclosed distinctly Russell bodies with different sizes and shapes. In conclusion, 4 weeks aged turkeys developed lymphoid tissue in the harderian gland which is a critical age for vaccination in this avian species.

A 62-Year-Old African American Woman With Stable Lymphadenopathy Over 5 Years

Abstract The patient is a 62-year-old African American female with medical history of Plummer Vinson syndrome and CT scan in 2014 showing lymphadenopathy of the porta hepatis. Needle core biopsy of enlarged lymph nodes revealed sheets of polytypic plasma cells with no aberrant B or T cells or granulomas or hyaline-like depositions identified. The findings were suspicious for unicentric Castleman disease. In 2019, the patient presented with chronic fatigue without other systemic symptoms. Contrast CT revealed persistent retroperitoneal lymphadenopathy and calcified granulomas in the liver, spleen, and lung. Excision of portal and celiac nodes showed preserved nodal architecture with mild mantle zone expansion and marked hyaline-like deposition in the follicles. Sheets of mature plasma cells and focally increased vasculatures were seen in the interfollicular areas. Focally frequent nonnecrotizing granulomas were noted. Occasional giant cells and rare asteroid bodies were identified in the granulomas. Immunohistochemistry stain workup revealed normal distribution of the B and T lymphocytes. CD138 stain highlighted marked increased plasma cells and HHV8 was negative. Immunofluorescence microscopy for kappa and lambda did not show monoclonality; IgA was positive in <10% of plasma cells; special stain for Congo red and serum amyloid associated protein (SSA) IHC stain were strongly positive in the follicles. The overall findings are consistent with reactive lymphadenopathy with interfollicular polytypic plasmacytosis, noncaseating granuloma formation, and reactive amyloidosis. The features of mantle zone hyperplasia and polytypic plasmacytosis are suggestive of HHV8-negative/idiopathic unicentric Castleman disease. This case possibly represents an example of concurrent amyloidosis, sarcoidosis, and idiopathic unicentric Castleman disease. Given that SSA was recently reported to have a pathogenic role in granulomatous inflammation of sarcoidosis through sustained inflammatory reaction, the possibility of increased SSA triggering sarcoidoisis in this patient can be considered. However, the exact pathogenesis of these concurrent findings needs further investigation and clarification.

Use of injectable platelet‐rich fibrin in the treatment of plasma cell mucositis of the oral cavity refractory to corticosteroid therapy: A case report

Plasma cell mucositis (PCM) is a rare benign disease affecting adults characterized by an erythematous mucosa, an epithelial hyperplasia, and a dense submucosal infiltration of mainly mature plasma cells. PCM has been treated with topical, intralesional, and systemic corticosteroids, antibiotics, and topical cyclosporin with unreliable results and questionable benefits. Here, we present a case of PCM, refractory to previous treatments, treated with i-PRF (injectable platelet-rich fibrin) injections. The infiltrations were performed once a week for 2 months. There were no adverse reactions to the treatment. The pain gradually reduced until the score of zero at the fourth infiltration, and the patient remained free of pain during the whole study period. Clinically, we did not obtain a complete healing of the lesion, but a reduced perilesional inflammatory infiltrate was observed at a distance. Therefore, we can conclude that i-PRF has been effective in the management of pain in PMC but does not result in complete healing of the disease.

Abstract 3101: Functional analysis of HDAC11 in plasma cell development and multiple myeloma survival

Abstract Though HDAC11 expression is confirmed in B cells and plasma cells (PC), its functions in these cells remain largely unknown. In this study, we attempted a functional analysis of HDAC11 in PC development along with its pro-tumorigenic function in multiple myeloma (MM) cells. Mouse models, including a transgenic mouse strain expressing eGFP under the regulation of the HDAC11 promoter (Tg-HDAC11-eGFP), and also an HDA11-deficient mouse (B6.HDAC11-/-) were studied to establish the importance of HDAC11 in PC biology. Pharmacologic inhibition of HDAC11 in MM cell lines was accomplished by using elevenostat (ES), a new HDAC11-selective inhibitor. Post-translational modifications and subcellular localization changes induced by ES exposure were assessed by western blotting of fractionated cell lysates, while co-immunoprecipitation (Co-IP) and proximity ligation assays (in situ PLA) were used to identify a binding partner for HDAC11. Studies in Tg-HDAC11-eGFP mice revealed that HDAC11 expression in B cell lymphopoiesis was minimally detectable prior to B cell activation but demonstrated strong induction upon maturation into a PC. Consistent with this, PC development was markedly impaired in the absence of HDAC11. Furthermore, HDAC11-selective inhibitor ES showed significant cytotoxic potential, measured by CCK-8 assay, in different MM cell lines and primary MM cells that express moderate to high level of HDAC11, with IC50 values ranging 0.6-2.0 µM. Apoptotic cell death was confirmed via detection of activated caspase-3 and annexin/propidium iodide staining by flow cytometry. Consistently, MM cell lines expressing null/very low level of HDAC11 were insensitive to ES. Moreover, combining ES with proteasome inhibitors bortezomib and carfilzomib resulted in significant synergistic effects. Furthermore, ES-treated cells showed a time-dependent alteration in the subcellular localization of HDAC11, it gradually disappeared from the nuclear fractions with simultaneous upregulation in cytoplasmic fractions. Inhibition of HDAC11 also caused downstream suppression of several pro-tumorigenic factors in MM cells, including IRF4 and c-Myc. Moreover, a novel interaction between HDAC11 and IRF4, an essential regulator of PC differentiation and MM survival, was identified by using PLA and Co-IP. HDAC11 dynamically interacts with IRF4 which can be induced by LPS and IL-6 stimulation and inhibited by ES, indicating the involvement of HDAC11 in the IRF4-mediated regulatory circuit. We also identified changes in the extent of IRF4 acetylation upon genomic or pharmacological induction/inhibition of HDAC11. Overall, we observe that targeted inhibition of HDAC11 can impair MM cell survival and overcome acquired resistance to proteasome inhibitors. Furthermore, we identify IRF4 as a nuclear binding partner of HDAC11 and propose this interaction as a candidate mechanism regulating PC maturation and survival. Citation Format: A G M MOSTOFA, Jason Brayer. Functional analysis of HDAC11 in plasma cell development and multiple myeloma survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3101.

PLASMA CELL GINGIVITIS DUE TO COSMETICS RELATED IODOPROPYNYL BUTYLCARBAMATE (IPBC) ALLERGY IN A TEENAGE FEMALE PATIENT MASKING AS DESQUAMATIVE GINGIVITIS.

Plasma cell gingivitis (PCG) is a rare lesion found on the attached and free gingiva, often extending to the mucogingival junction. Clinically, PCG can appear as sharply delineated erythematous lesions which can be accompanied by edema. We present a case of PCG in a 13 year old female patient which clinically presented as generalized desquamative gingivitis involving the facial aspects of the attached and marginal gingiva that persisted despite substituting to a non flavored dentifrice and failure to elicit any suspected drug or food allergies. Dermatologic patch testing proved positive for Iodopropynyl Butylcarbamate (IPBC), a water based preservative or biocide used in personal care products. Microscopically, there was an intense inflammatory infiltrate in the lamina propria composed predominantly of mature plasma cells. Immunohistochemistry and in situ hybridization showed marked unrestricted cytoplasmic positivity for kappa and lambda light chains. IPBC is used in personal care products comprising lip balms, moisturizers, sunscreens, concealers and body washes due to its effectiveness at preventing fungal growth in topical products. The maximum level for safe use in leave on products is 0.1% but cosmetic products continue to use 10 times more than the safe levels. In the differential spectrum it is pertinent to discriminate erosive lichen planus, cicatricial pemphigoid, acute leukemia, HIV infection clinically; multiple myeloma and plasmacytoma histologically. Our case highlights the importance of patch testing for IPBC allergies in the oral mucosa. IPBC can lead to sensitization and contact dermatitis due to prolonged exposure; as its use in cosmetics continues to rise, and it is difficult to completely eliminate exposure to products containing IPBC especially in the context of teenaged girls and adult female patients.

PF560 A MATURATION INDEX DEFINES NEWLY DIAGNOSED MULTIPLE MYELOMA PATIENTS WITH ADVANCED IMMUNOPHENOTYPIC AND MOLECULAR DIFFERENTIATION PROFILES ASSOCIATED WITH POOR PROGNOSIS

Background:The differentiation status plasticity of Multiple Myeloma (MM) plasma cells (PC) is an adaptive strategy that might confer a specific fitness to tumour cells, enabling their interaction to an evolving microenvironment. Therefore, MM clones are not constantly “fit”, and the immunophenotypic profile of the fittest MM PCs might be the expression of specific genetic and genomic programs, emerging under therapeutic pressure and promoting tumour development. However, the genomic background that supports any diverse plasma cell differentiation phenotypes has not yet been inferred.Aims:To correlate the genetic and genomic background with the immunophenotypic profile of MM clones at diagnosis, in order to stratify patients (pts) according to a Maturation Index, and ultimately to evaluate the impact of this stratification on the disease outcome.Methods:114 newly diagnosed MM pts were included in the study. For each pts, both the neoplastic PCs and CD19+ B cells compartments were characterized by 6‐color multi‐parameter flow cytometry analysis, combining CD138‐PE, CD38‐PE‐Cy7, CD56‐APC‐Cy7, CD20‐APC, CD19‐FITC, CD27‐APC, CD45‐PerCp, CD28‐APC, CD117‐FITC, CD28‐APC, CD81‐APC, IgK‐APC and IgL‐FITC. Both whole‐genome copy number alterations (CNAs) and a 25‐genes targeted mutational panel were assessed in CD138+ PCs. A custom ddPCR assay was employed to evaluate the self‐renewal status of PCs.Results:In order to define a Maturation Index, pts were evaluated for: a) differential expression of CD19/CD81 markers; b) level of chromosomal instability (CIN); c) self‐renewal status.According to the CD19 and CD81 markers co‐expression, we were able to stratify pts in 3 different subgroups, recapitulating a progressive PC maturation process: the most immature, which included pts with PCs CD19+/CD81+ (20/114 = 17%); an intermediate CD19‐/CD81+ phenotype subgroup (40/114 = 35%); and the CD19‐/CD81‐ PC subgroup (54/114 = 48%), whose clone was mainly composed by most mature plasma cells.CIN was defined according to both the total CNAs count and the portion of genome changed (GC). Pts with an advanced differentiation status (CD19‐/CD81‐) were more frequently associated to a high CIN (medium tot. CNAs = 550, % GC ≥ 25%), including a higher prevalence of high‐risk features. Indeed, 1p deletion (FAF1), 16q deletion (WWOX, FANCA) and 17p deletion (TP53) were the most recurrent abnormalities. Genomic instability was also confirmed by a higher incidence of clonal pathogenic mutations in critical genes (e.g. NRAS, KRAS, TP53). Interestingly, the application of a 10 Hh‐genes signature, resuming the Hedgehog pathway, demonstrated that PCs with more advanced differentiation status displayed a substantial overexpression of all the genes, indicating a more proliferative, aggressive and, possibly, persistent phenotype.Finally, the presence of a more mature PCs characterized pts carrying baseline clinical features associated to bad prognosis (e.g. n. PET lesions, k/l ratio, ISS III, β2‐microglobulin; p < .05). In addition, these pts tended to obtain high quality response rates (≥VGPR) to PI induction therapy.Summary/Conclusion:An iMMature index defined pts with an advanced differentiation status both at immunophenotypic as well as at molecular level, and this is lastly associated with a prevalence of bad prognosis features. Chromosomal instability, together with cellular phenotypic plasticity, represents an important, yet poorly defined, mechanism by which MM clones accelerate their own evolution and survival.Acknowledgements: AIRC IG2014, Fondazione Berlucchi.

Bone Marrow Plasma Cells Modulate Local Myeloid-Lineage Differentiation via IL-10.

Bone marrow plasma cells have been reported to represent a major source of IL-10; however, the impact of plasma cell derived IL-10 in that tissue remains poorly understood. We confirm in this study that even in the absence of acute immune reactions, mature plasma cells represent the dominant IL-10+ cell population in the bone marrow, and identify myeloid-lineage cells as a main local target for plasma cell derived IL-10. Using Vert-X IL-10 transcriptional reporter mice, we found that more than 50% of all IL-10+ cells in bone marrow were CD138+ plasma cells, while other IL-10+ B lineage cells were nearly absent in this organ. Accordingly, IL-10 was found in the supernatants of short-term cultures of FACS-sorted bone marrow plasma cells, confirming IL-10 secretion from these cells. IL-10+ bone marrow plasma cells showed a B220−/CD19−/MHCII low phenotype suggesting that these cells represent a mature differentiation stage. Approximately 5% of bone marrow leucocytes expressed the IL-10 receptor (IL-10R), most of them being CD115+/Ly6C+/CD11c− monocytes. Compared to littermate controls, young B lineage specific IL-10 KO mice showed increased numbers of CD115+ cells but normal populations of other myeloid cell types in bone marrow. However, at 7 months of age B lineage specific IL-10 KO mice exhibited increased populations of CD115+ myeloid and CD11c+ dendritic cells (DCs), and showed reduced F4/80 expression in this tissue; hence, indicating that bone marrow plasma cells modulate the differentiation of local myeloid lineage cells via IL-10, and that this effect increases with age. The effects of B cell/plasma cell derived IL-10 on the differentiation of CD115+, CD11c+, and F4/80+ myeloid cells were confirmed in co-culture experiments. Together, these data support the idea that IL-10 production is not limited to early plasma cell stages in peripheral tissues but is also an important feature of mature plasma cells in the bone marrow. Moreover, we provide evidence that already under homeostatic conditions in the absence of acute immune reactions, bone marrow plasma cells represent a non-redundant source for IL-10 that modulates local myeloid lineage differentiation. This is particularly relevant in older individuals.

Inhibiting EZH2 function prevents and treats chronic graft vs host disease (cGVHD) by disrupting germinal center formation and plasma cell maturation

Abstract To determine whether targeting EZH2, a histone methyltransferase necessary for germinal center (GC) formation, could reduce murine cGVHD (cGVHD), we first used a clinically relevant murine model of cGVHD, which recapitulates multi-organ cGVHD with bronchiolitis obliterans (BOS). Bone marrow (BM) from EZH2 fl/fl CD19 Cre B6 mice and WT T cells, or WT B6 BM and EZH2 fl/fl CD4 Cre B6 T cells were given to conditioned B10.BR mice. EZH2 expression in BM and T cells was obligatory for cGVHD. To determine whether EZH2 function could be targeted to treat cGVHD, we tested a novel EZH2 small molecule inhibitor, JQ5 with a favorable toxicity and efficacy profile to other inhibitors. Mice with established cGVHD given JQ5 had dramatically improved cGVHD. Similar results were seen in a sclerodermatous cGVHD model. To assess possible causes of this improvement we performed RNA-seq and ATAC-seq data of MACS purified CD19+ B cells obtained from control and JQ5 treated BOS cGVHD mice. Unsupervised gene set enrichment analysis and weighted gene correlation network analysis showed that among gene clusters associated with JQ5 treatment there was an enrichment for genes expressed in naïve B cells compared to plasma cells (GSEA4142) and an impairment of genes associated with late stage GC B cells compared to early follicular B cells (GSEA11961). This suggested that JQ5 impaired GC B cell and plasma cell maturation. Consistent with these data, splenic follicular cells, GC size and number, and lung resident PCs were reduced compared to cGVHD controls. Taken together these results show that EZH2 is necessary for the initiation of cGVHD and show for the first time that EZH2 inhibition using a novel inhibitor JQ5 can reverse established cGVHD in mice with BOS or scleroderma.