CD4+Foxp3−IL-10+Tr1 Cells Promote Relapse of Diffuse Large B Cell Lymphoma by Enhancing the Survival of Malignant B Cells and Suppressing Antitumor T Cell Immunity

Abstract Abstract 790 Several investigators have defined gene expression profiling (GEP) signatures in Diffuse Large B-cell Lymphoma (DLBCL) enriched for macrophage and stromal genes, suggesting an active innate immune response against lymphoma. We hypothesize that the malignant B-cells drive tumor-associated macrophage (TAM) dysfunction in a subset of patients with DLBCL which is relevant for their biology and prognosis. We performed GEP on TAM from diagnostic DLBCL in order to recognise key genes and pathways open to functional validation. Single cell suspensions from 8 DLBCL and 8 reactive lymph nodes were used in this study. TAMs were flow sorted using CD36 expression. cDNA synthesis and amplification was performed using the Nugen Ovation Pico WTA system and the Affymetrix GeneChip human gene 1.0 ST platform was used. We used bioinformatics analysis of GEP data from DLBCL whole tumors, DLBCL purified B-cells, in vitro manipulated macrophages and other immune cells in order to define macrophage-enriched genes as well as specific M1/M2 signatures. We identified a 221 gene signature that significantly distinguished DLBCL TAMs from control macrophages, with 165 genes upregulated and 56 genes downregulated. Moreover, a comparative transcriptome analysis of 22 diverse immune cell phenotypes/activation states (IRIS: GSE22886) revealed that 26% of these genes are highly macrophage-enriched. Gene Ontology analysis revealed an over-representation for transcripts involved in inflammatory response (p 6.8×10−23), wound healing (p 1.9×10−20), chemotaxis (p 3.2 ×10−9), and cell motility (p 1.7×10−7). Upregulated genes in TAMs included well known M1 (complement components, CXCL9 or CXCL10) as well as M2 genes (MSR, CD163 or MARCO) (Table 1). In our signature, there was enrichment for M1 compared to M2 genes as defined by bioinformatics analysis. TAMs showed overexpression of the CSF1R gene as well as the chemokines CCL2 and CCL5, suggesting an autocrine feed-back loop of macrophage chemotaxis and survival in DLBCL. Moreover, TAMs showed upregulation of the lymphocyte attractants CCL20, CXCL9 and CXCL10, together with T-cell immunosupressants indoleamine 2,3-dioxygenase 1 and PD-L1, which would support a role for macrophages in T-cell recruitment and dysfunction in DLBCL. We also saw strong upregulation of 7 metallothionein isoforms in TAMs. These are proteins known to be expressed in macrophages and linked to response to oxidative damage, modulation of inflammation and cell proliferation. However their role in cancer microenvironment is unclear. We describe for the first time the GEP from DLBCL TAMs. The TAM transcriptome has partial overlapping genes with both M1 and M2 gene signatures, but also has a characteristic GEP potentially driven by their presence in the DLBCL microenvironment. Although further molecular and functional validation is required, this data provides a platform of genes which serve as excellent candidates for future exploration to understand DLBCL pathogenesis and to define new therapeutic targets. Table 1: Genes of particular interest represented in our TAM signature. Probe collapse was done using the highest expression. The Benjamini-Hochberg multiple hypothesis test was used to determine significance (FDR 0.05). Gene IDs Gene description Log2 Fold Change Adjusted p value MT1E-H, L, M, X, MT2A metallothionein 1E-H, 1M, 1X, 2A 2.3 – 4.4 0.00145 - 0.00017 C1QA, B and C complement component 1, q subcomponent, A, B and C chains 3.1 – 3.6 0.00136 - 0.00255 C2 complement component 2 3.2 0.00136 CCL2, 4, 5, 8, 20 chemokine (C-C motif) ligand 2, 4, 5, 8 and 20 2.4 - 3.7 0.046 - 0.00130 CD163 CD163 molecule 4.6 0.00025 CD14 CD14 molecule 3.4 0.00091 CD274 CD274 molecule 3.7 0.00447 CSF1R colony stimulating factor 1 receptor 2.1 0.00886 CXCL9-11 chemokine (C-X-C motif) ligand 9, 10 and 11 4.2 – 4.4 0.012 - 0.00158 FCGR1B Fc fragment of IgG, high affinity Ib, receptor (CD64) 4.9 0.00091 FCGR2A Fc fragment of IgG, low affinity IIa, receptor (CD32) 3.3 0.00139 FCGR3A Fc fragment of IgG, low affinity IIIa, receptor (CD16a) 5.2 0.00011 IDO1 indoleamine 2,3-dioxygenase 1 4.8 0.00232 MARCO macrophage receptor with collagenous structure 2.6 0.02029 MSR1 macrophage scavenger receptor 1 3.7 0.01467 Disclosures: Gribben: Celgene: Honoraria; Roche: Honoraria; Pharmacyclics: Honoraria; GSK: Honoraria; Mundipharma: Honoraria; Gilead: Honoraria.

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Spatial Signatures Identify Immune Escape via PD-1 as a Defining Feature of T-cell/Histiocyte-rich Large B-cell Lymphoma

Blood ◽

10.1182/blood.2020006464 ◽

2020 ◽

Cited By ~ 1

Author(s):

Gabriel K. Griffin ◽

Jason L. Weirather ◽

Margaretha Roemer ◽

Mikel Lipschitz ◽

Alyssa Kelley ◽

...

Keyword(s):

T Cell ◽

B Cells ◽

B Cell ◽

Immune Escape ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Spatially Resolved ◽

Large B Cell Lymphoma ◽

Immune Signatures ◽

Large B Cell

T-cell/histiocyte-rich large B cell lymphoma (TCRLBCL) is an aggressive variant of diffuse large B cell lymphoma (DLBCL) characterized by rare malignant B cells within a robust but ineffective immune cell infiltrate. The mechanistic basis of immune escape in TCRLBCL is poorly defined and not targeted therapeutically. We performed a genetic and quantitative spatial analysis of the PD-1/PD-L1 pathway in a multi-institutional cohort of TCRLBCLs and found that malignant B cells harbor PD-L1/PD-L2 copy gain or amplification in 64% of cases, which is associated with increased PD-L1 expression (p = 0.0111). By directed and unsupervised spatial analyses of multi-parametric cell phenotypic data within the tumor microenvironment, we found that TCRLBCL is characterized by tumor-immune 'neighborhoods' in which malignant B cells are surrounded by exceptionally high numbers of PD-L1-expressing TAMs and PD-1-positive T cells. Further, unbiased clustering of spatially-resolved immune signatures distinguished TCRLBCL from related subtypes of B-cell lymphoma, including classic Hodgkin lymphoma (cHL) and DLBCL-NOS. Finally, we observed clinical responses to PD-1 blockade in three of five patients with relapsed/refractory TCRLBCL who were enrolled in clinical trials for refractory hematologic malignancies, including two complete responses and one partial response. Taken together, these data implicate PD-1 signaling as an immune escape pathway in TCRLBCL, and also support the potential utility of spatially-resolved immune signatures to aid the diagnostic classification and immunotherapeutic prioritization of diverse tumor types.

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New Molecular and Therapeutic Insights into Canine Diffuse Large B Cell Lymphoma Elucidates the Role of the Dog As a Model for Human Disease

Blood ◽

10.1182/blood-2018-99-112763 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 4173-4173

Author(s):

Luca Aresu ◽

Serena Ferraresso ◽

Laura Marconato ◽

Luciano Cascione ◽

Sara Napoli ◽

...

Keyword(s):

T Cell ◽

B Cells ◽

Clinical Outcome ◽

B Cell ◽

Cell Line ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Rna Seq ◽

Large B Cell Lymphoma ◽

Large B Cell

Abstract Background. Diffuse large B-cell lymphoma (DLBCL) is the commonest lymphoma in both humans and dogs. Canine DLBCL (cDLBCL) is considered an ideal comparative model for drug development, but a complete genomic characterization of this tumor is still lacking. In this study, we report an integrated analysis to comprehensively define the molecular mechanisms of cDLBCL and possible associations with clinical outcome. Methods. Fifty cDLBCLs were analyzed by RNA-Seq, methyl-CpG-binding sequencing and array comparative genomic hybridization. Normal B-cells derived from lymph nodes of 11 healthy dogs were used as controls.Additionally, immunohistochemistry, in vitroand in vivoexperiments were performed as validation analyses. Results.Compared to normal B-cells, cDLBCL showed a marked up-regulation of genes involved in the PI3K/mTOR and NF-κB pathways, including several TLRs in association with MYD88, indicating mechanisms similar to the human activated B cell-like subtype DLBCL. Both RNA-Seq and methylation sequencing led to the identification of two groups of cDLBCLs bearing different clinical outcome. The two groups did not overlap with the human germinal center B-cell (GCB) and the activated B-cell-like (ABC) DLBCL subtypes or the human DLBCL consensus clusters. The dogs with the poorest outcome presented a signature largely defined by markers of T-cell-mediated immune responses, with a high expression of PDL-1, PD-1 and CTLA-4, also validated in an independent cohort of cDLBCL by immunohistochemistry. These data provide a strong rationale for the use of cDLBCL to study immune checkpoint modulators. The observed high expression of PI3K/mTOR pathway genes was confirmed and validated achieving a clear anti-tumor activity with the use of the PI3K-delta inhibitor idelalisib and of the novel dual PI3K/mTOR inhibitor bimiralisib in the cDLBCL cell line CLBL-1. The cDLBCLs showed an up-regulation of MYC and of its targets, sustained by recurrent gains in the chromosome 13, where the oncogene is located, in approximately half of the cases. Thus, we have exposed the cDLBCL cell line CLBL-1 to the BET inhibitor birabresib (OTX015) and to the BRD4 degrader MZ1. Both compounds caused a significant reduction in the proliferation of tumor cells, and this effect was stronger especially with the second compound. Exposure to MZ1 determined an important downregulation of MYC and also of LIN28B, the most overexpressed transcript in cDLBCL when compared to controls. While LIN28B does not seem to be a relevant gene for human DLBCL, its overexpression causes murine T-cell lymphomas (Beachy et al, Blood 2011), and there is a direct association of MYC with LIN28B promoter resulting in transcriptional transactivation (Chang et al, PNAS 2009). Here, LIN28B genetic silencing in the CLBL-1 lead to a reduction in cell growth, opening new therapeutic target perspectives in canine lymphoma. Conclusions. We have reported the first large next generation sequencing study investigating the cDLBCL transcriptome, methylome and the genome-wide CNVs. We identified deregulated pathways and individual transcripts providing therapeutic targets, including an immune-related signature affecting the outcome of a subgroup of cDLBCL. Our data sustain the use of cDLBCL as comparative models for human DLBCL but also highlight differences that must be kept in consideration. Disclosures Hillmann: PIQUR Therapeutics AG: Employment. Wymann:PIQUR Therapeutics AG: Employment, Equity Ownership, Patents & Royalties.

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

Journal of the Endocrine Society ◽

10.1210/jendso/bvab048.1796 ◽

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.

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T Cell Histiocyte Rich Large B Cell Lymphoma Presenting as Hemophagocytic Lymphohistiocytosis: An Uncommon Presentation of a Rare Disease

Case Reports in Oncological Medicine ◽

10.1155/2017/6428461 ◽

2017 ◽

Vol 2017 ◽

pp. 1-5

Author(s):

Uroosa Ibrahim ◽

Gwenalyn Garcia ◽

Amina Saqib ◽

Shafinaz Hussein ◽

Qun Dai

Keyword(s):

T Cell ◽

B Cells ◽

B Cell ◽

Rare Disease ◽

Hemophagocytic Lymphohistiocytosis ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Biologically Active ◽

Large B Cell Lymphoma ◽

Large B Cell

T cell histiocyte rich large B cell lymphoma (THRLBCL) is a rare subtype of non-Hodgkin’s lymphoma characterized by malignant B cells with reactive T lymphocytes. The pathophysiology is thought to involve cytokine-mediated evasion of T cell immune response by malignant B cells. It usually presents at an advanced stage with extranodal involvement. An extremely unusual manifestation of the disease is hemophagocytic lymphohistiocytosis (HLH) which is a hyperinflammatory disorder. We present a case of a 43-year-old male who presented with recurrent fever and recent radiologic imaging showing splenomegaly and right inguinal lymphadenopathy. On presentation, he had a fever of 105°F. Laboratory work-up was consistent with pancytopenia, elevated lactate dehydrogenase, elevated D-dimer, and a ferritin of 24,247 ng/mL. The patient was started on steroid therapy. An excisional biopsy of the right inguinal lymph node was consistent with a diagnosis of THRLBCL and the patient subsequently received six cycles of chemotherapy with R-CHOP (Rituximab, Cyclophosphamide, Doxorubicin, Vincristine, and Prednisone) after which a PET-CT scan showed no evidence of biologically active disease and ferritin was down to 822 ng/mL. We discuss the clinical manifestations and diagnostic and therapeutic considerations of this rare disease along with a review of reported cases in the literature.

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CD4+ T cell-mediated cytotoxicity is associated with MHC class II expression on malignant CD19+ B cells in diffuse large B cell lymphoma

Experimental Cell Research ◽

10.1016/j.yexcr.2017.11.029 ◽

2018 ◽

Vol 362 (2) ◽

pp. 287-292 ◽

Cited By ~ 2

Author(s):

Yong Zhou ◽

Jie Zha ◽

Zhijuan Lin ◽

Zhihong Fang ◽

Hanyan Zeng ◽

...

Keyword(s):

T Cell ◽

B Cells ◽

B Cell ◽

Mhc Class Ii ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Class Ii ◽

Cd4 T Cell ◽

Large B Cell Lymphoma ◽

Large B Cell

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Long-Term Survival and Gradual Recovery of B Cells in Patients (Pts) with Refractory Large B Cell Lymphoma (LBCL) Treated with Axicabtagene Ciloleucel (Axi-Cel)

Transplantation and Cellular Therapy ◽

10.1016/s2666-6367(21)00520-0 ◽

2021 ◽

Vol 27 (3) ◽

pp. S404-S405

Author(s):

Caron A. Jacobson ◽

Frederick L. Locke ◽

Armin Ghobadi ◽

David B. Miklos ◽

Lazaros J. Lekakis ◽

...

Keyword(s):

B Cells ◽

B Cell ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Term Survival ◽

Long Term Survival ◽

Large B Cell Lymphoma ◽

Large B Cell

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Use of immune repertoire sequencing to resolve discordant microscopic and immunochemical findings in a case of T cell-rich large B cell lymphoma in a young dog

BMC Veterinary Research ◽

10.1186/s12917-021-02783-3 ◽

2021 ◽

Vol 17 (1) ◽

Author(s):

Gary Kwok Cheong Lee ◽

Dorothee Bienzle ◽

Stefan Matthias Keller ◽

Mei-Hua Hwang ◽

Nikos Darzentas ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cell ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Immune Repertoire ◽

Large B Cell Lymphoma ◽

Repertoire Sequencing ◽

Reactive Lymphocytes ◽

Large B Cell

Abstract Background Lymphocytic neoplasms with frequent reactive lymphocytes are uncommonly reported in dogs, and can pose a diagnostic challenge. Different diagnostic modalities such as cytology, flow cytometry, histopathology, immunohistochemistry, and clonality testing, are sometimes required for a diagnosis. This report illustrates the value of using a multi-modal diagnostic approach to decipher a complex lymphocytic tumor, and introduces immune repertoire sequencing as a diagnostic adjunct. Case presentation A 10-month-old Great Dane was referred for marked ascites. Cytologic analysis of abdominal fluid and hepatic aspirates revealed a mixed lymphocyte population including numerous large lymphocytes, yielding a diagnosis of lymphoma. Flow cytometrically, abdominal fluid lymphocytes were highly positive for CD4, CD5, CD18, CD45, and MHC II, consistent with T cell lymphoma. Due to a rapidly deteriorating clinical condition, the dog was euthanized. Post mortem histologic evaluation showed effacement of the liver by aggregates of B cells surrounded by T cells, suggestive of hepatic T cell-rich large B cell lymphoma. Immune repertoire sequencing confirmed the presence of clonal B cells in the liver but not the abdominal fluid, whereas reactive T cells with shared, polyclonal immune repertoires were found in both locations. Conclusions T cell-rich large B cell lymphoma is a rare neoplasm in dogs that may be challenging to diagnose and classify due to mixed lymphocyte populations. In this case, the results of histopathology, immunohistochemistry and immune repertoire sequencing were most consistent with a hepatic B cell neoplasm and reactive T cells exfoliating into the abdominal fluid. Immune repertoire sequencing was helpful in delineating neoplastic from reactive lymphocytes and characterizing repertoire overlap in both compartments. The potential pitfalls of equating atypical cytomorphology and monotypic marker expression in neoplasia are highlighted.

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