scholarly journals Angioimmunoblastic T-Cell Lymphoma: Molecular Characterization of Clonal T and B-Cells and a Patient Derived Xenograft Model of Coexisting T and B-Cell Proliferations

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1572-1572
Author(s):  
Xiaoxian Zhao ◽  
Bartlomiej Przychodzen ◽  
Juraj Bodo ◽  
Lisa Durkin ◽  
Daniel Lindner ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Tumor Cells ◽  
Cell Lymphoma ◽  
T And B Cells ◽  
Pdx Model ◽  
Cell Proliferations

Introduction: Angioimmunoblastic T-cell lymphoma (AITL) is a rare and aggressive type of lymphoma that accounts for about 20% of peripheral T-cell lymphomas with a 5 year overall survival rate of 30%. As most patients relapse after anthracycline-containing regimens and newer agents such as histone deacetylase inhibitors, other novel therapeutic approaches are needed. Signaling lymphocytic activation molecule F7 (SLAMF7), a molecule expressed on a subset of T-cells, activated B cells and myeloma cells, is an attractive target to explore based on our previous studies showing SLAMF7 expression in a subset of AITL cases. The association of AITL with Epstein Barr virus (EBV) positive B-cells is nearly always present and the efficacy of treatment in such patients with significant EBV viral load is not well-understood. In this study, we performed the molecular characterization of an aggressive EBV+ AITL case, established a patient-derived xenograft (PDX) AITL model of coexisting T and B-cell proliferations and evaluated novel therapeutic strategies. Methods: Primary tumor cells were injected into a NSG mouse. Flow cytometry, immunohistochemistry (IHC), CISH-EBER and BIOMED 2 PCR based clonality studies were used to confirm the engraftment and compared the consistency of engrafted tumor cells with the primary sample. Genomic DNA extracted from sorted T and B cells and from paired normal neutrophils of the original patient were subjected to Whole Exome sequencing (WES). In vivo AITL PDX model trials were tested for the efficacy of romidepsin (Rom), elotuzumab (Elo), rituximab (Rit) and in combinations of these drugs. Results: A 53 year old woman with AITL was treated with 6 cycles of CHOEP followed by autologous stem cell transplantation. 3 months after transplantation (9 months after diagnosis) she developed progressive fatigue and arthralgias. PET-CT scan showed new cervical, thoracic, abdominal and pelvic lymphadenopathy. A cervical lymph node biopsy was performed to confirm relapse. IHC staining showed atypical T cells expressing CD2, CD3, CD4, CD5, CD7, CD10, BCL6, PD1, SLAMF7 and CXCL13. Scattered CD20+/EBER+ B-immunoblasts were present with focal large clusters/small sheets. Primary tumor cells engrafted in NSG mouse via tail vein injection caused splenomegaly. Flow cytometry assay demonstrated the engraftment of tumor cells in peripheral blood, bone marrow and spleen tissue. CD3+CD19- cells dominated the engrafted cells in all three tissues. Histologic examination and immunophenotyping (IHC and EBER staining) of spleen were consistent with primary tumor tissue. Engrafted tumor cells were capable of serial passage in NSG mice with an increasing malignant B cell percentage that mimics the situation in which the B-cell component masks an underlying T-cell lymphoma in humans. T-cell receptor gene rearrangement assay confirmed the clonal identity of engrafted T-cells matched the primary relapsed tumor. A clonal IGH rearrangement of engrafted B-cells was also detected, while no monoclonal B-cell population was detected in the relapsed AITL sample, possibly due to the low number of EBV+ B-cells present in that biopsy. WES of sorted malignant T-cells showed 33 mutants in 31genes, including RhoA G17V, TET2,STAT3 and VAV1, previously described in AITL or other T-cell lymphomas. In parallel WES assay, 9 mutations were found in 9 genes from sorted EBV+ B immunoblasts. A PDX model using cells harvested from the second passage showed single agent, Elo or Rit, extended the survival of mice compared to the control group (p < 0.05). Rom alone had no such effect (p = 0.27). Combination of Rit with either Elo or Rom further improved survival compared to each single agent exposed cohort (p < 0.05). There was no significant difference between Rit/Elo and Rit/Rom (p = 0.067). PARP cleavage by IHC was higher in the Rit/Rom and Rit/Elo groups compared to other cohorts. Expression of SLAMF7 in a subset of engrafted T and B cells of the control mouse were confirmed via flow cytometry assay. Conclusions: To date, this is the first molecular characterization of AITL tumor cells in comparison with associated EBV+ B cells and use of such a PDX model for therapeutic evaluation of agents targeting both malignant T and B cells simultaneously. The in vivo data support further clinical investigation of applying elotuzumab or romidepsin in combination with rituximab in AITL containing EBV-positive B-cell proliferations. Disclosures Maciejewski: Novartis: Consultancy; Alexion: Consultancy. Hsi:Abbvie: Research Funding; Eli Lilly: Research Funding; Jazz: Consultancy; Cleveland Clinic&Abbvie Biotherapeutics Inc: Patents & Royalties: US8,603,477 B2.

Nodular lymphocyte-predominant Hodgkin lymphoma with nodules resembling T-cell/histiocyte-rich B-cell lymphoma: differential diagnosis between nodular lymphocyte-predominant Hodgkin lymphoma and T-cell/histiocyte-rich B-cell lymphoma

Blood ◽  
2003 ◽  
Vol 102 (10) ◽  
pp. 3753-3758 ◽  
Author(s):  
Ludmila Boudová ◽  
Emina Torlakovic ◽  
Jan Delabie ◽  
Peter Reimer ◽  
Beate Pfistner ◽  
...  
Keyword(s):  
T Cells ◽  
Differential Diagnosis ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Hodgkin Lymphoma ◽  
Tumor Cells ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
World Health

AbstractNodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) and T-cell/histiocyte-rich B-cell lymphoma (T/HRBCL) are distinct tumors and are treated differently. They are linked by a morphologic and probably a biologic continuum, which renders the differential diagnosis difficult. To develop criteria to distinguish the entities along the morphologic continuum, we correlated the lymph node architecture and immunophenotype of both tumor cells and reactive components of 235 neoplasms in the spectrum of NLPHL and T/HRBCL with clinical data. Two hundred and eighteen cases fitted the World Health Organization (WHO) criteria of NLPHL (139) or T/HRBCL (79). While tumor cells in both entities were immunophenotypically similar, background composition differed: in NLPHL small B cells and CD3+CD4+CD57+ T cells were common, whereas in T/HRBCL, CD8+ cytotoxic T cells and histiocytes dominated. Follicular dendritic cells (FDCs) formed expanded meshworks in NLPHL, whereas they were absent in T/HRBCL. Seventeen cases represented a gray zone: within FDC meshworks, neoplastic B cells resided in a background depleted of small B cells but rich in T cells and histiocytes. Tumor cells either were loosely scattered or formed clusters, thus resembling areas of either T/HRBCL or inflammatory diffuse large BCL (DLBCL) within the nodules. Patients with these NLPHLs with T-cell/histiocyte-rich nodules presented at a high stage and with B symptoms, as in T/HRBCL, but had an excellent survival, as in NLPHL. This morphologic pattern suggests a biologic continuum between NLPHL and T/HRBCL. (Blood. 2003;102:3753-3758)

Tumor Microenvironment In Nodular Lymphocyte Predominant Hodgkin Lymphoma (NLPHL) Influences Occurrence of Relapses and Progression to Large Cell Lymphoma

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2684-2684
Author(s):  
Nasir Bakshi ◽  
Mansoor Aljabry ◽  
Saad Akhter ◽  
Irfan Maghfoor ◽  
Ayman Mashi
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Tumor Microenvironment ◽  
B Cell ◽  
Hodgkin Lymphoma ◽  
Clinical Course ◽  
Cell Lymphoma ◽  
Lp Cells

Abstract Abstract 2684 NLPHL accounts for 6.5% of all Hodgkin lymphoma cases in the West. It is characterized by a nodular or a nodular & diffuse proliferation of scattered large atypical CD20+ neoplastic B-cells referred to as lymphocyte predominant (LP) cells and typically associated with small lymphocytes mainly of B-cell type. Patients with NLPHL typically have an indolent clinical course but can frequently relapse. Progression to a higher grade lymphoma, notably T-cell/Histiocyte rich B-cell lymphoma (T/HRBCL) has been described in a relatively small number of cases. Because of its rarity, limited information is available about the role of non-neoplastic lymphocytes in NLPHL. Some studies suggest that NLPHL with T-cell rich background may behave differently than the conventional type with predominance of B-cells within the nodules. The purpose of this study was to evaluate outcomes of differential tumor microenvironment namely B-cell versus T-cell rich in patients with NLPHL. We document the clinicopathologic profiles of 29 patients with biopsy proven NLPHL, consisting of 22 male & 7 female, median age 26 years (range, 13–80 years). All patients had lymphoadenopathy & 2 cases showed extranodal involvement in addition to nodal disease. Two patients had a bulky mass, and three had stage 4 disease at presentation. The pathological diagnoses was reviewed and confirmed by an expert hematopathologist in all 29 cases. The LP cells in all cases had a prototypic immunophenotype of CD20+, CD79a+, PU.1+, Bcl-6+, CD15− CD30− & Fascin−. T/HRBCL was excluded as all cases demonstrated preservation of follicular dendritic meshwork by CD21 staining. The meshwork was expanded in 20 cases & in 9 cases it was partially disrupted evincing an irregular architectural pattern. Epstein-Barr Virus encoded RNA by in situ hybridization was negative in 8/8 cases tested. 27/29 patients received systemic multi-agent chemotherapy consisting of: doxorubicin, bleomycin, vinblastine, and dacarbacin (ABVD), 24 patients; cyclophosphamide, doxorubicin, vincristin, and prednisone (CHOP), 2 patients; Rituximab + CHOP (R-CHOP), 1 patient. 9/29 (31%) cases underwent autologous stem cell transplant. One patient in stage 2A refused therapy and one patient (stage 3A) developed significantly decreased cardiac ejection fraction following initial 2 cycles of ABVD. Both of these cases did not have adequate follow-up information available. Results: Twelve of the 29 cases (42%) were designated as having T-cell rich background population, whereas 17 (58%) were considered as conventional variant with a vast predominance of non-neoplastic small lymphocytes being B-cells. A few of the cases seemed to show admixture of both B-cells & T-cells. Comparing T-cell rich & B-cell rich background NLPHL no significant differences were detected in clinical parameters: age, sex, and stage at presentation, absolute lymphocyte count, LDH & Hb. All 27 (100%) patients in this study responded to first-line treatment: 23 with complete response & 4 with partial response. 13/27 (48%) had relapse/s. Five cases had more than one relapses. No patient died within a clinical follow-up period ranging from 18 to 84 months. When the overall survival (OS) of T-cell rich NLPHL was compared with the conventional variant there was no statistical significance between the two groups (log rank p= 0.1206). However, comparison of relapse rate showed that cases with T-cell rich background had higher relapse rate as well as greater incidence of multiple relapses as compared to B-cell rich type of NLPHL even after adjusting for the type of treatment received (log rank p= 0.003). Moreover, 2/12 (17%) T-cell rich NLPHL cases showed transformation to a high grade lymphoma (both T/HRBCL) at the time of recurrence. These findings suggest that in NLPHL a tumor microenvironment rich in T-cells rather than B-cells is characterized by an unfavorable clinical course although OS appears to be similar. These cases perhaps represent a distinctive clinicopathologic variant within the framework of NLPHL. Lately, the term ‘NLPHL with nodules resembling T/HRBCL’ has been used to express the immunobiological overlap between these two entities. It is possible that such cases could be regarded as “intermediate lymphomas” treading between NLPHL and T/HRLBCL. Further studies using gene array profiling analysis may help clarify the molecular differences between these closely related entities. Disclosures: No relevant conflicts of interest to declare.

A New Platform For Trivalent Bispecific Antibodies Used For T-Cell Redirected Killing Of B-Cell Malignancies

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3078-3078
Author(s):  
Diane L Rossi ◽  
Edmund A Rossi ◽  
David M Goldenberg ◽  
Chien-Hsing Chang
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Cell Surface ◽  
B Cell ◽  
Tumor Cells ◽  
Stock Option ◽  
B Cell Malignancies ◽  
Hla Dr ◽  
Close Proximity

Abstract Background Various formats of bispecific antibodies (bsAbs) to redirect effector T cells for the targeted killing of tumor cells have shown considerable promise both pre-clinically and clinically. The scFv-based constructs, including BiTE and DART, which bind monovalently to CD3 on T cells and to the target antigen on tumor cells, exhibit fast blood clearance and neurological toxicity due to their small size (∼55 kDa). Herein, we describe the generation of novel T-cell redirecting trivalent bsAbs comprising an anti-CD3 scFv covalently conjugated to a stabilized F(ab)2. The design was initially characterized with a prototype construct designated (19)-3s, which specifically targets CD19 on B cells. A panel of trivalent bsAbs was evaluated for their potential use in targeted T-cell immunotherapy of various B-cell malignancies. Potential advantages of this design include bivalent binding to tumor cells, a larger size (∼130 kDa) to preclude rapid renal clearance and penetration of the blood-brain barrier, and potent T-cell mediated cytotoxicity. Methods The DOCK-AND-LOCKTM (DNLTM) method was used to generate a panel of B-cell targeting bsAbs, (19)-3s, (20)-3s, (22)-3s, and (C2)-3s, which target CD19, CD20, CD22, and HLA-DR, respectively. This was achieved by combining a stabilized anti-X F(ab)2 with an anti-CD3-scFv, resulting in a homogeneous covalent structure of the designed composition, as shown by LC-MS, SE-HPLC, ELISA, SDS-PAGE, and immunoblot analyses. Each construct can mediate the formation of immunological synapses between T cells and malignant B cells, resulting in T-cell activation. At an E:T ratio of 10:1, using isolated T cells as effector cells, the bsAbs induced potent T-cell-mediated cytotoxicity in various B-cell malignancies, including Burkitt lymphomas (Daudi, Ramos, Namalwa), mantle cell lymphoma (Jeko-1), and acute lymphoblastic leukemia (Nalm-6). A non-tumor binding control, (14)-3s, induced only moderate T-cell killing at >10 nM. The nature of the antigen/epitope, particularly its size and proximity to the cell surface, appears to be more important than antigen density for T-cell retargeting potency (Table 1). It is likely that (20)-3s is consistently more potent than (19)-3s and (C2)-3s, even when the expression of CD19 or HLA-DR is considerably higher than CD20, as seen with Namalwa and Jeko-1, respectively. This is likely because the CD20 epitope comprises a small extracellular loop having close proximity to the cell surface. When compared directly using Daudi, (22)-3s was the least potent. Compared to CD19 and CD20, CD22 is expressed at the lowest density, is a rapidly internalizing antigen, and its epitope is further away from the cell surface; each of these factors may contribute to its reduced potency. Finally, sensitivity to T-cell retargeted killing is cell-line-dependent, as observed using (19)-3s, where Raji (IC50 >3 nM) is largely unresponsive yet Ramos (IC50 = 2 pM) is highly sensitive, even though the former expresses higher CD19 antigen density. Conclusions (19)-3s, (20)-3s, (22)-3s, and (C2)-3s can bind T cells and target B cells simultaneously and induce T-cell-mediated killing in vitro. The modular nature of the DNL method allowed the rapid production of several related conjugates for redirected T-cell killing of various B-cell malignancies, without the need for additional recombinant engineering and protein production. The close proximity of the CD20 extracellular epitope to the cell surface results in the highest potency for (20)-3s, which is an attractive candidate bsAb for use in this platform. We are currently evaluating the in vivo activity of these constructs to determine if this novel bsAb format offers additional advantages. Disclosures: Rossi: Immunomedics, Inc.: Employment. Rossi:Immunomedics, Inc.: Employment. Goldenberg:Immunomedics: Employment, stock options, stock options Patents & Royalties. Chang:Immunomedics, Inc: Employment, Stock option Other; IBC Pharmaceuticals, Inc.: Employment, Stock option, Stock option Other.

Constitutive Chemokine Receptor Expression in B-Cell Non-Hodgkin’s Lymphoma.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3590-3590
Author(s):  
Michelle S. Bryson ◽  
Ruth F. Jarrett ◽  
Lesley Sheild ◽  
Gerard J. Graham
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Mantle Cell Lymphoma ◽  
Hodgkin’S Lymphoma ◽  
Memory T Cells ◽  
Cell Lymphoma ◽  
Hodgkin's Lymphoma ◽  
Mantle Cell

Abstract Chemokines are small peptides (∼8-14KDa) that play an essential role in both the innate and adaptive immune system. Chemokines are primarily involved in leukocyte trafficking, but are also involved in a number of cellular mechanisms. They elicit their effect through G-protein coupled receptors, the chemokine receptors (CKR). Functionally chemokines and their receptors are classified as inflammatory or constitutive. Constitutive CKRs and their ligands have a role in numerous diseases including malignancy, chronic inflammation and HIV infection. This study aimed to examine constitutive CKR expression in sub-types of B-cell NHL, of which there are limited studies so far. Lymph node preparations from patients with NHL were examined by flow cytometry using antibodies to CD20, CCR4, CCR6, CCR7, CCR9, CCR10, CXCR4 and CXCR5. The percentage of CD20 positive cells expressing the CKR under investigation was then calculated. The following cases were examined; follicular lymphoma (FL), n=11, Diffuse large B-cell lymphoma (DLBCL), n=11, mantle cell lymphoma (MCL) n=17, Burkitt’s lymphoma (BL), n=9 and MALT lymphoma, n=10. A number of differences between NHL sub-types were detected. FL cases generally had a lower expression of all the CKRs. CXCR5 and CXCR4 expression was high in all sub-types (>84% of B-cells) with no significant differences found, this would be expected as these CKRs are widely expressed in all B-cells. CCR10 expression was low or absent, with no significant differences detected. CCR6 and CCR9 show highest expression in MALT lymphomas, consistent with previous studies, but in comparison with other sub-types the differences was not significant. The most significant results were found with CCR7 and CCR4. CCR7 is expressed on naive T-cells, memory T-cells, B-cells and dendritic cells and is involved in the homing of lymphocytes to lymph nodes. CCR7 is currently the second most commonly reported CKR to be upregulated in malignancy, after CXCR4 and is related. We found very high levels of CCR7 in Mantle cell lymphoma (>90% of B-cells) as compared to other sub-types (p=0.005). CCR4 is expressed on Th2 and Treg lymphocytes, memory T cells and in a small subset of mature B-cells. CCR4 expression in T-cells has been correlated with an adverse prognosis in T-cell NHL and Hodgkin’s lymphoma, yet no systematic studies looking at CCR4 expression in B-cell neoplasms has been reported. These results showed a significant increase in CCR4 expression (>50% of B-cells) in DLBCL, MCL, MALT and BL as compared to FL (p<0.0001). We showed that there are differences in constitutive CKR expression in the different B-cell NHL types, with CCR4 expression being the most interesting finding. How CCR4 expression relates to prognosis in these lymphomas is as yet unknown but is under investigation. Targeting of the chemokine system using anti-CCR4 is already being used in clinical trials for T-cell neoplasms, and may be of potential benefit in selected B-cell neoplasms. Furthermore, the development of anti-CCR7 strategies may prove to be of benefit in the traditionally poor prognosis MCL patients.

scholarly journals A Rare Case of T-Cell Histiocyte Rich Large B-Cell Lymphoma of the Thyroid in a Patient With Hashimotos

2021 ◽  
Vol 5 (Supplement_1) ◽  
pp. A879-A880
Author(s):  
Abir Zainal ◽  
Jhansi Maradana ◽  
Mira Torres
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Thyroid Gland ◽  
Lymph Nodes ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Large B Cell Lymphoma ◽  
Large B Cell

Abstract Introduction: T-cell/histiocyte-rich large B-cell lymphoma (THRLBCL) is a rare form of large B-cell lymphoma, which usually involves the lymph nodes exclusively. We describe a patient with Hashimoto’s thyroiditis who was discovered to have THRLBCL arising from the thyroid. Clinical Case: A 78-year-old female with a history of Hashimoto’s thyroiditis noted increase in the size of her left thyroid lobe for two months despite normal TSH on Levothyroxine, prompting an ultrasound which revealed several enlarged left sided cervical lymph nodes and an enlarged left thyroid gland. Cytology from an FNA of a left level 3 lymph node showed atypical lymphoid infiltrate featuring scattered large atypical cells in a background of small lymphocytes. Immunohistochemical testing was PAX5+, CD30- and CD15-. Cytology from an FNA of left thyroid revealed identical changes and immunohistochemistry demonstrated PAX5+ and CD20+. Concurrent flow cytometric studies demonstrated increased CD4 to CD8 ratio among T cells. Excisional biopsy of a left cervical lymph node confirmed a diagnosis of THRLBCL. PET/CT exhibited lymphadenopathy above her diaphragm and splenic involvement. Her bone marrow biopsy was negative for involvement. She was deemed Stage III with international prognostic index (IPI) of 2 corresponding with low-intermediate risk. She was commenced on chemotherapy R-CHOP with plan to complete 6 cycles. Discussion: THRLBCL is characterized by scattered atypical B lymphocytes on a background of T lymphocytes and histiocytes. Usually, T-cells are predominantly CD8+, in contrast to our patient. Some studies identified cases of predominant CD4+ and PD1+ T cells. Cytology revealed scattered small B-cells and large B-cells, a feature that is not typically seen in THRLBCL. A diagnosis of diffuse transformation of nodular lymphocyte predominant Hodgkin lymphoma was considered but the diffuse proliferation outside of CD21+ and involvement of the thyroid is not compatible with such diagnosis. Similarly, a diagnosis of follicular helper T-cell lymphoma with admixed large B-cells was considered but while PD1+ CD4+ T cells are present, there was no aberrant antigen expression by flow cytometry or T cell clonality. THRLBCL mainly involves lymph nodes and presents at advanced Ann Arbor stages with high IPI. Malignant lymphomas of the thyroid gland are exceedingly rare, accounting for 2% of thyroid cancers, out of which the literature reveals a single case report of THRLBCL arising from the thyroid. THRLBCL represents an aggressive form of lymphoma and is treated according to stage-matched DLBCL, although the effects of Rituximab in this population is variable. Conclusion: Hashimoto’s is considered a risk for thyroid lymphoma usually diffuse large B-cell lymphoma and MALT lymphoma. We present a rare case of THRLBCL occurring in the setting of Hashimoto’s with acute thyroid gland enlargement.

scholarly journals MiR-17-92 Is Required for the Pathogenicity of T and B Cells in Chronic Gvhd

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4535-4535
Author(s):  
Yongxia Wu ◽  
Steven D Schutt ◽  
Ryan P Flynn ◽  
Mengmeng Zhang ◽  
Hung D Nguyen ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Antibody Production ◽  
Cd4 T Cell ◽  
B Cell Differentiation ◽  
T And B Cells ◽  
And Function

Abstract Chronic graft-versus-host disease (cGVHD) remains to be a major cause of mortality and morbidity after allogeneic hematopoietic cell transplantation (allo-HCT). cGVHD is characterized as autoimmune-like fibrosis and antibody production, mediated by pathogenic T and B cells. Through producing pro-inflammatory cytokines, CD4 T cells are the driving force of cGVHD. Donor B cells augment the pathogenesis of cGVHD not only by acting as antigen-presenting cells (APCs) and promoting CD4 T-cell expansion and survival, but also by producing autoantibodies. microRNA (miR)-17-92 has been shown to regulate T-cell immunity including allogeneic, anti-viral, and anti-tumor responses. Recently, miR-17-92 was found to act together with Bcl-6 to promote the differentiation of Follicular help T (Tfh) cells. Furthermore, B-cell deficiency of miR-17-92 impairs IgG2c production. Since Tfh differentiation and antibody production are required for the development of cGVHD, we hypothesize that miR-17-92 contributes to the pathogenesis of cGVHD by promoting pathogenic T- and B-cell responses. By using Cre-loxp system, we generated B6 mice with conditional deficiency of miR-17-92 in T cells (CD4cre), B cells (CD19cre), or both (CD4CD19cre). aGVHD to cGVHD transition model (B6 to BALB/c) was utilized to test the effects of individual and combinational deficiency of miR-17-92 in T and/or B cells in the development of cGVHD. BALB/c mice were lethally irradiated and transferred with splenocytes plus BM cells derived from CD4cre, CD19cre or CD4CD19cre miR-17-92flox/flox B6 mice. WT B6 (Cre- miR-17-92flox/flox) mice were used as control donors. A significantly reduction of GVHD mortality was observed only in the recipients with CD4CD19cre grafts, but not with CD4cre or CD19cre grafts. Deficiency of miR-17-92 in donor T or B cells indeed improved the clinical manifestation of cGVHD, but the deficiency in both T and B cells showed further improvement, indicating the additive role of miR-17-92 in T and B cells in the pathogenesis of cGVHD. Mechanistically, deficiency of miR-17-92 in T cells resulted in the reduction of Tfh generation (Fig. A), germinal center (GC) B-cell and plasma cell differentiation, and the expression of MHC-II and CD86 on donor B cells in recipient spleens. Furthermore, deficiency of miR-17-92 in B cells significantly reduced the levels of total IgG and IgG2c in recipient serum (Fig. A). These data suggest that miR-17-92 contributes to both T- and B-cell differentiation and function, which is required for the development of cGVHD. To extend our findings, we used a bronchiolitis obliterans cGVHD model (B6 to B10.BR). Recipient mice were pre-conditioned and received either BM alone from WT or CD19cre B6 mice, or BM plus purified T cells from WT or CD4cre B6 mice. Deficiency of miR-17-92 in T cells or BM-derived B cells resulted in significant improvement in pulmonary functions in recipient mice, as demonstrated by a decrease in resistance and elastance and an increase in compliance (Fig. B). Consistently, we found that miR-17-92 promoted Tfh and GC B-cell differentiation (Fig. B), while inhibiting differentiation of T follicular regulatory cells in recipient spleens 60 days after allo-HCT. For translational purpose, we tested whether inhibition of miR-17-92 could ameliorate cGVHD using locked nucleic acid (LNA) antagomirs specific for miR-17 or miR-19, key members in this microRNA cluster. In a SLE cGVHD model (DBA2 to BALB/c), administration of anti-miR-17, but not anti-miR-19, significantly suppressed the incidence of proteinuria and the severity of clinical manifestation by inhibiting donor splenocyte expansion, expression of costimulatory molecules on donor B cells, and differentiation of GC B cells and plasma cells (Fig. C). In addition, systemic delivery of anti-miR-17 significantly improved skin cGVHD by restraining IL-17 producing CD4 T-cell infiltration in skin-draining lymph nodes in a scleroderma-cGVHD model (B10.D2 to BALB/c). Taken together, the current work reveals that miR-17-92 is required for T- and B-cell differentiation and function, and thus for the development of cGVHD. Furthermore, pharmacological inhibition of miR-17 represents a potential therapeutic strategy for the control of cGVHD after allo-HCT. Figure Figure. Disclosures No relevant conflicts of interest to declare.

TCL1 Expression in B-Cell Tumors Is Dynamically Regulated by Maturation Stage, Proliferation, and the Follicular Microenvironment.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4333-4333
Author(s):  
Marco Herling ◽  
Ryuji Kobayashi ◽  
Kaushali A. Patel ◽  
Kong Chao Chang ◽  
Ellen Schlette ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Tumor Cells ◽  
Germinal Center ◽  
Cell Lymphoma ◽  
Maturation Stage ◽  
Dynamic Regulation ◽  
B Cell Subsets ◽  
Human B Cell

Abstract The kinase comodulator TCL1 is the primary initiating oncogene in T-cell prolymphocytic leukemia and can produce B-cell or T-cell chronic lymphocytic leukemia (CLL) following transgenic expression in mice. Given its strong expression in some non-neoplastic B-cell subsets, the role of TCL1 as an oncogene in human B-cell tumors is less clear. Using a recently developed TCL1 monoclonal antibody (clone 1–21), we examined the relationship between TCL1 expression and B-cell maturation stage in tumor tissue arrays, lymphoma cell lines, primary tumor samples and in vitro stimulation assays. Results were compared with immunohistochemical expression of a variety of maturation, activation and cell proliferation markers and with the somatic hypermutation status determined by VH transcript analysis (<2% sequence divergence in VH regarded as “pre-germinal center”). In non-neoplastic B-cells, TCL1 was strongly and uniformly expressed in naive B-cell subsets, but was variably expressed in subsets of germinal center B-cells, with complete absence of expression in immunoblasts and plasma cells. In germinal center B-cells, TCL1 was expressed more strongly in the quiescent centrocyte fraction than in the proliferating centroblasts. This pattern was replicated in human B-cell lymphoma lines and tumors with complete absence of TCL1 in myeloma cases (n = 35) and monocytoid B-cells in marginal zone lymphoma (n = 8) and dim or absent expression in the majority of the mutated/post-germinal center subset of B-CLL (10/15, 67%). In contrast, TCL1 showed strong but modulated expression in the majority of unmutated/pre-germinal center type of B-CLL (14/19, 74%), and mantle cell lymphoma (MCL, 44/57, 84%), and less frequently in follicular lymphoma (FL, 21/47, 45%). In B-CLL, TCL1 was overexpressed in non-proliferating cells within the pseudo-follicular proliferation centers but was markedly downregulated in the CD23+bright PCNA+ proliferating tumor cell component. Similarly in MCL, TCL1 was downregulated in the proliferative component but upregulated in tumor cells in follicular and mantle zone locations versus the diffuse areas. In FL, the highest levels of TCL1 expression were found in those cases that most strongly expressed the germinal center markers CD10 and bcl-6, but TCL1 staining was inversely correlated with expression of proliferation and activation markers, such as CD23. In 3 FL cases with multiple biopsies at different tissue sites, TCL1 was downregulated in tumor cells at extranodal sites as compared to those within lymph node follicles. Thus, the dynamic regulation of TCL1 in B-cells and derived tumors is likely due to changes in the balance between stimulatory microenvironmental influences within the lymphoid follicle and inhibitory signals during cell cycle progression/proliferation. In post-germinal center tumors, TCL1 expression is effectively silenced, and no longer exhibits this dynamic regulation pattern. These studies strongly suggest that TCL1 exerts its effects in promoting cell survival in quiescent B-cell subsets prior to and in the absence of an (antigenic) proliferative signal.

Targeting DKK1 for the Immunotherapy of B-Cell Lymphomas.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 465-465
Author(s):  
Jianfei Qian ◽  
Sungyoul Hong ◽  
Liang Zhang ◽  
Yuhuan Zheng ◽  
Haiyan Li ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Tumor Cells ◽  
Cell Lines ◽  
Cd8 T Cells ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Lymphoma Cells ◽  
Specific Ctls

Abstract Abstract 465 Immunotherapy may complement the current treatments for lymphomas. The lack of suitable shared lymphoma-associated antigens limits its applicability. Therefore, identification and utilization of novel and more potent tumor-associated antigens, particularly those shared among patients, are urgently needed to improve the efficacy of immunotherapy in the diseases. Recent studies have shown that Dickkopf-1 (DKK1), a secreted protein and Wnt signaling pathway inhibitor, is highly expressed by myeloma and other tumor cells, and is absent from normal tissues and organs except placenta and prostate. In the present study we demonstrated that DKK1 is also overexpressed in mantle cell lymphoma (MCL) and diffuse large B-cell lymphoma (DLBCL). Using DKK1 peptide-pulsed dendritic cells (DCs), we successfully generated HLA-A*0201+ DKK1-specific CTL lines and clones in vitro. These CTLs effectively lysed DKK1+/HLA-A*0201+ lymphoma cell lines Jeko-1 and Granta 519 cells, but not DKK1-/HLA-A*0201+ BJAB, RL and Mino cells nor DKK1+/HLA-A*020- CA46 and Daudi cells. Furthermore, the T-cell clones efficiently killed DKK1+/HLA-A*0201+ primary B-cell lymphoma cells from patients but not lymphoma cells from DKK1–/HLA-A*0201+ patients. HLA-ABC or HLA-A*0201 blocking mAbs significantly inhibited T cell-mediated cytotoxicity against peptide-pulsed T2 cells (P < .01, compared with medium control). No inhibitory effect was observed with mAb against HLA-DR and isotype control IgG. The results indicate that the cytotoxicity was attributed to MHC class I and more specifically, HLA-A*0201-restricted CD8+ CTLs. The CTLs did not kill DKK1–/HLA-A*0201+ DCs, B cells, or PBMCs, These results suggest that the CTLs recognized DKK1 peptides that are naturally processed and presented in the context of HLA-A*0201 molecules on lymphoma cells. To determine the in vivo antitumor activity, NOD-SCID and SCID-hu mice were used for lymphoma cell lines and primary lymphoma cells, respectively. Mice were treated with DKK1-specific CTLs after tumor established in NOD-SCID and SCID-hu mice. Control mice were treated with naïve CD8+ T cells or PBS alone. Tumor burden was measured according to levels of circulating human B2M, and survival rates were determined. Low levels (< 50 ng/ml) of circulating human B2M were detected in group treated DKK1-specific CTLs, while high levels (≥ 150 ng/ml) of circulating human B2M were detected in control mice. In SCID-hu model, X-ray examination showed that established tumors were eradicated in 60% mice treated with DKK1-specific CTLs, while large tumor burdens were found in all control mice. In NOD-SCID model, 40% of mice survived with the treatment of DKK1-specific CTLs. TUNEL assay further confirmed that tumor cells were lysed by DKK1-specific CTLs not naïve CD8+ T cells. These results indicate that DKK1-specific CTLs are able to eradicate established, patient-derived primary B- cell lymphoma in the hosts and adoptive transfer of DKK1-specific CTLs may be used for B-cell lymphoma therapy. Disclosures: No relevant conflicts of interest to declare.

Examining the Heterogeneity of Follicular Lymphoma By Multi-Parameter Flow Cyotmetry in Previously Untreated Patients

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2947-2947
Author(s):  
Debra K Czerwinski ◽  
Steven R Long ◽  
Michael Khodadoust ◽  
Matthew J. Frank ◽  
Adel Kardosh ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Follicular Lymphoma ◽  
B Cell ◽  
Tumor Cells ◽  
Immune Cells ◽  
Research Funding ◽  
T Cell Exhaustion ◽  
Cell Exhaustion

Abstract BACKGROUND: Follicular lymphoma (FL) is an indolent form of Non-Hodgkin B cell lymphoma that remains incurable with present therapies. Derived from germinal center B cells, FL B cells experience ongoing hypermutation of the immunoglobulin variable region gene. In addition, Michael Green, et al (PNAS; 2015), reported the presence of numerous somatic mutations to include those of the chromatin-modifying genes. These mutations accumulate over the course of the disease and play an important role in regulating gene transcription, B cell development and immune interactions. Furthermore, FL tumors maintain a resemblance to primary lymphoid follicles, and as such, present with a number of infiltrating immune cells, especially T cells, the numbers of which vary from patient to patient. The close association and interaction of these immune cells with the tumor B cells play an important part in determining the disease biology (Dave SS, et al. N Engl J Med; 2004). For instance, tumor B cells, through cell-cell contact with these immune cells and/or through secretion of inhibitory cytokines such as TGF-b and IL-10, induce T cell exhaustion and apoptosis as well as suppressive T cell phenotypes (FoxP3+ T Regulatory cells) thus evading immune eradication (Yang Z-Z, et al. Blood 2007 and Ai WZ, et al. IntJ Cancer; 2009). They also promote their own survival and proliferation through their interaction with resident T follicular helper cells via CD40L/CD40 interactions (Ame'-Thomas P, et al. Blood; 2005). As a corollary to an ongoing clinical trial, we received fine needle aspirates (FNAs) of easily accessible tumors from 14 patients with FL prior to any treatment. 6 of these patients had samples taken from a second site simultaneously. All samples were processed within 24 hours into a single-cell suspension; red blood cells were lysed. Cells were then stained with antibodies to delineate T, B, NK, dendritic, and myeloid cells, as well as their subsets. Antibodies against activation antigens, T cell exhaustion, inhibition and function were also used to characterize these cells. Finally, the cells were run on a 17-parameter LSRII (Becton Dickinson) and data analyzed via Cytobank, a web-based data storage and analysis tool. PURPOSE: To better understand the biology of FL as represented by protein expression by the tumor cells and the immune cells that make up the microenvironment. We will especially look to evaluate the heterogeneity inherent in FL by flow cytometry across patients as well as within any one individual. RESULTS: Each sample is stained with 4 panels of antibodies, 13 antibodies each, allowing us to measure over 100 cell subsets. A quick preview of all data shows that there is a high variability between patients in the percentage of T cells within the microenvironment (37.7% + 16.6% of all cells collected from all samples). This variability is represented by the differences in the CD4 T cell compartment (27.6 + 12.9%) and to a lesser degree in the CD8 compartment (7.7 + 3.7%). To note, this variability in T cells does not correlate with time from diagnosis to sample collection which ranged from 3.4 years to approximately 5 months. Also, this is in contrast to the similar percentage of CD4 and CD8 T cells expressing PD-1 (55.5 + 8.8% and 46.0 + 8.9%, respectively) across patients. Notably, there is much less variability from site to site within each patient then between patients as demonstrated by Figure 1 where Site A and Site B are 2 separate lesions within each patient listed, sampled at the same time. Since FL presumably begins in a single site in the body and then becomes disseminated, the fact that a characteristic relationship exists between tumor cells and immune cells wherever the disease is found implies a mutual interdependence of the tumor cells in each case and their immune host component. CONCLUSION: Follicular lymphoma is a very heterogeneous disease as would be expected by the diversity of mutations seen at the genomic level. This heterogeneity is also apparent in the microenvironment from one patient to another. Conversely, different tumor sites within each patient have a characteristic and fixed relationship to their immune microenvironment. The emergence of novel therapies for FL, including checkpoint antibodies such as anti-PD-1 and anti-PD-L1 and small molecules such as Ibrutinib, will be informed by understanding the differences as well as the similarities in each case of FL. Disclosures Levy: Kite Pharma: Consultancy; Five Prime Therapeutics: Consultancy; Innate Pharma: Consultancy; Beigene: Consultancy; Corvus: Consultancy; Dynavax: Research Funding; Pharmacyclics: Research Funding.

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