Immunohistochemical staining of Bcl2, cMyc, Ki67, and CD3 in DLBCL subtype representative tumor samples (same as Supplementary Figure S1). Note that AD091967 is positive for both Bcl2 and cMyc. Cases AB189717, AD550620, and AE575816 show high proliferation rates (80%, 80%, 60%) Scale bars represent 50 µm.

Hematoxylin and eosin (H&E), CD20, CD10, Bcl6, and MUM1 stainings in 4 non-GCB and 2 GCB cases representative for each DLBCL subtype. Bars, 50 µm.

AbstractPurpose: The aurora A kinase inhibitor alisertib demonstrated single-agent clinical activity and preclinical synergy with vincristine/rituximab in B-cell non-Hodgkin lymphoma (B-NHL). This phase I study aimed to determine the safety and recommended phase II dose (RP2D) of alisertib in combination with rituximab ± vincristine in patients with relapsed/refractory aggressive B-NHL. Patients and Methods: Patients with relapsed/refractory, diffuse, large, or other aggressive B-NHL received oral alisertib 50 mg b.i.d. days 1 to 7, plus i.v. rituximab 375 mg/m2 on day 1, for up to eight 21-day cycles (MR). Patients in subsequent cohorts (3 + 3 design) received increasing doses of alisertib (30 mg starting dose; 10 mg increments) b.i.d. days 1 to 7 plus rituximab and vincristine [1.4 mg/m2 (maximum 2 mg) days 1, 8] for 8 cycles (MRV). Patients benefiting could continue single-agent alisertib beyond 8 cycles. Cell-of-origin and MYC/BCL2 IHC was performed on available archival tissue. Results: Forty-five patients participated. The alisertib RP2D for MR was 50 mg b.i.d. For MRV (n = 32), the RP2D was determined as 40 mg b.i.d. [1 dose-limiting toxicity (DLT) at 40 mg; 2 DLTs at 50 mg]. Drug-related adverse events were reported in 89% of patients, the most common was neutropenia (47%). Seven patients had complete responses (CR), 7 had partial responses (PRs); 9 of 20 (45%) patients at the MRV RP2D responded (4 CRs, 5 PRs), all with non–germinal center B-cell (GCB) diffuse large B-cell lymphoma (DLBCL). Conclusions: The combination of alisertib 50 mg b.i.d. plus rituximab or alisertib 40 mg b.i.d. plus rituximab and vincristine was well tolerated and demonstrated activity in non-GCB DLBCL.

Agents targeting the PD1-PDL1 axis have transformed cancer therapy. Factors that influence clinical response to PD1-PDL1 inhibitors include tumor mutational burden, immune infiltration of the tumor, and local PDL1 expression. To identify peripheral correlates of the anti-tumor immune response in the absence of checkpoint blockade, we performed a retrospective study of circulating T cell subpopulations and matched tumor gene expression in melanoma and non-small cell lung cancer (NSCLC) patients. Notably, both melanoma and NSCLC patients whose tumors exhibited increased inflammatory gene transcripts presented high CD4(+) and CD8(+) central memory T cell (CM) to effector T cell (Eff) ratios in blood. Consequently, we evaluated CM/Eff T cell ratios in a second cohort of NSCLC. The data showed that high CM/Eff T cell ratios correlated with increased tumor PDL1 expression. Furthermore, of the 22 patients within this NSCLC cohort who received nivolumab, those with high CM/Eff T cell ratios, had longer progression-free survival (PFS) (median survival: 91 vs. 215 days). These findings show that by providing a window into the state of the immune system, peripheral T cell subpopulations inform about the state of the anti-tumor immune response and identify potential blood biomarkers of clinical response to checkpoint inhibitors in melanoma and NSCLC.

Background: The unprecedented success of immuno-oncology (I-O) agents targeting the cytotoxic T lymphocyte-associated antigen 4 and programmed death-1/programmed death-ligand 1 pathways has stimulated the rapid development of other I-O agents against novel immune targets. Bristol-Myers Squibb has designed a novel phase II platform trial, the Fast Real-time Assessment of Combination Therapies in Immuno-ONcology (FRACTION) Program, to efficiently identify promising combinations for patients with specific malignancies. The concept and study design of the FRACTION Program-currently ongoing in patients with advanced non-small-cell lung cancer (FRACTION-Lung), gastric cancer (FRACTION-Gastric Cancer) and renal cell carcinoma (FRACTION-RCC)-are described. Methods: The FRACTION Program comprises open-label, phase II studies that use adaptive randomisation designs with rolling combination regimens. Master Protocols provide the overall study design framework, whereas Sub-Protocols introduced over time provide details on specific I-O combination therapies to which patients may be randomised. In a Master Protocol, patients are enrolled into different Study Tracks based on characteristics such as prior I-O therapy experience. Patients who progress may be rerandomised to other combination regimens from any ongoing Sub-Protocol. Primary objectives are to assess objective response rate, median duration of response and progression-free survival rate at 24 weeks; the secondary objective is to investigate safety and tolerability. Biomarker collection before and on treatment will facilitate identification of patient subsets who benefit most from each therapy. Conclusions: The FRACTION Program allows for the evaluation of multiple I-O combinations through individual studies for specific tumours using an adaptive trial design and continuous enrolment. (C) 2018 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

To discern features of non-Hodgkin lymphomas (NHL) that are autonomous from those that are shaped by the tumor environment (TE), we used patient-derived xenografts (PDX) to probe the effects on neoplastic cells of manipulating the TE. Properties of neoplastic cells that are often considered to be autonomous include their relative independence from stromal support, their relative survival and/or proliferation advantages compared with nonneoplastic cells, and their state of differentiation. Prior approaches to creation of PDX models likely select for neoplasms, which are the most capable of engraftment, potentially masking the effects of the TE. To overcome this bias, we developed a robust protocol that rapidly produced xenografts with more than 85% of unselected, cryo-preserved, B-cell NHL specimens, including low-grade tumors such as follicular and marginal zone lymphoma. To discern features that are shaped by the TE, we extensively studied 4 low-grade lymphoma specimens. B-cell engraftment required components of the native TE; specifically, CD4+ cells. The relative survival of neoplastic compared with nonneoplastic B cells was not autonomous in 2 specimens; specifically, neoplastic B cells from 2 specimens showed a greater dependence on the TE than normal B cells for engraftment. Furthermore, the differentiation of neoplastic B cells was dependent on the TE; mature B-cell neoplasms converted to plasmacytoma-like lesions in the grafts. These results highlight the central and patient-specific roles of the TE in maintaining the relative survival of neoplastic cells compared with normal cells and in controlling the differentiation of neoplastic cells.

Abstract Background: Immunotherapy with PD-1 inhibitors is associated with increased survival in NSCLC. While PD-L1 expression enriches for clinical benefit, additional predictive biomarkers are critically needed to further define patient subsets associated with specific response/resistance profiles and to develop optimal combination strategies. We hypothesized that integration of pathology features derived from H&E and PD-L1 immunohistochemistry with genetic/genomic data from NSCLC could identify phenotypic/genotypic associations that highlight specific immunosuppressive mechanisms. Towards that end, we present data from an initial pilot analysis. Methods: Twenty-nine NSCLC (23 adenocarcinoma; 6 squamous cell carcinoma) were included. H&E-stained sections were scored for tumor grade (1-3), relative proportion of tumor stroma (stroma score 1-3), percent necrosis and intensity of chronic inflammatory infiltrate (1-3+). PD-L1 immunohistochemistry was performed using the DAKO 28-8 antibody. PD-L1 expression was scored in tumor cells by modified H-score and by the predominant pattern observed - diffuse, heterogeneous, tumor-stroma interface or negative, and in immune cells using a semi-quantitative intensity scale (1-3+). DNA and RNA extracts from FFPE tissue were subjected to whole exome sequencing and RNAseq from which mutation load, oncogene/tumor suppressor genotypes and inflammation signatures were derived. Results: The highest PD-L1 H-scores were associated with diffuse expression patterns and were observed in high grade tumors (Grade 3). Deleterious mutations in TP53, STK11, KEAP1, KRAS, EGFR and MET occurred at expected frequencies. These mutations occurred across multiple PD-L1 expression patterns except for STK11 which was restricted to PD-L1 negative tumors (p=0.077, Fisher exact test). STK11-mutant tumors also displayed a lower PD-L1+ inflammation score (p=0.046, Fisher exact test). Trends toward an increase in mutation load with increasing levels of chronic inflammation on H&E stain as well as with PD-L1+ chronic immune infiltrates were noted. Inflammation signatures derived from RNAseq showed an association between diffuse PD-L1 expression by IHC and the highest levels of inflammation mRNA signatures comprising T-cells (CD8, Tregs), B-cells, macrophages and MDSCs. Conclusions: The potential association between tumor grade, PD-L1 expression, intensity of the immune infiltrate and mutation load raise the possibility that tumor morphology predicts mutation load and associated immune response. The finding that STK11 mutation is restricted to PD-L1 negative tumors suggests an immunosuppressive mechanism is invoked in this setting. This dataset is currently being expanded to establish the significance of these findings. Citation Format: Robin Edwards, Patrik Vitazka, Peter Szabo, Han Chang, John Cogswell, Hao Tang, Kaushal Desai, Darren Locke, Qiuyan Wu, Joseph Szustakowski, Steven Bernstein, Michele Cleary. Cross-platform integrative analysis of NSCLC reveals association between PD-L1 expression patterns and tumor genetic profile [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 593. doi:10.1158/1538-7445.AM2017-593

As we previously described, intra-peritoneal injection of single cell suspensions (including non-neoplastic cells) from >90% of primary B cell non-Hodgkin’s lymphoma specimens results in lymphoma cell growth in the omentum of the NSG mouse strain (NOD-scid IL2rg null) (ASH Meeting on Lymphoma Biology 2014; abstract #144). To date, we have shown reproducible engraftment of 17/20 specimens (including 9 follicular, 5 marginal zone, 4 diffuse large B cell, 2 mantle cell). Molecular markers specific for each patient’s tumor (including clone specific features of the IGK, IGH, BCL2 and BCL1 loci) provided verification that all 17 of the engrafted tumors contained the same clone identified in the original specimen. Each tumor was implanted and engrafted in at least 3 mice (range 3-41) and each had a characteristic and often distinctive time course of engraftment, propensity to disseminate, and histologic features, indicating that the behaviors of the xenografts reflected intrinsic properties of the original tumors.

We have previously shown that regulatory T cells (Tregs) infiltrating follicular lymphoma lymph nodes are quantitatively and qualitatively different than those infiltrating normal and reactive nodes. To gain insight into how such Treg populations differ, we performed RNA sequence (RNAseq) analyses on flow sorted Tregs from all three sources. We identify several molecules that could contribute to the observed increased suppressive capacity of follicular lymphoma nodal tregs, including upregulation of CTLA-4, IL-10, and GITR, all confirmed by protein expression. In addition, we identify, and confirm functionally, a novel mechanism by which Tregs target to and accumulate within a human tumor microenvironment, through the down regulation of S1PR1, SELL (L-selectin) and CCR7, potentially resulting in greater lymph node retention. In addition we identify and confirm functionally the upregulation of the chemokine receptor CXCR5 as well as the secretion of the chemokines CXCL13 and IL-16 demonstrating the unique ability of the follicular derived Tregs to localize and accumulate within not only the malignant lymph node, but also localize and accumulate within the malignant B cell follicle itself. Such findings offer significant new insights into how follicular lymphoma nodal Tregs may contribute to the biology of follicular lymphoma and identify several novel therapeutic targets.

Aggressive induction chemotherapy followed by autologous haematopoietic stem cell transplant (auto-HCT) is effective for younger patients with mantle cell lymphoma (MCL). However, the optimal induction regimen is widely debated. The Southwestern Oncology Group S1106 trial was designed to assess rituximab plus hyperCVAD/MTX/ARAC (hyperfractionated cyclophosphamide, vincristine, doxorubicin and dexamethasone, alternating with high dose cytarabine and methotrexate) (RH) versus rituximab plus bendamustine (RB) in a randomized phase II trial to select a pre-transplant induction regimen for future development. Patients had previously untreated stage III, IV, or bulky stage II MCL and received either 4 cycles of RH or 6 cycles of RB, followed by auto-HCT. Fifty-three of a planned 160 patients were accrued; an unacceptably high mobilization failure rate (29%) on the RH arm prompted premature study closure. The estimated 2-year progression-free survival (PFS) was 81% vs. 82% and overall survival (OS) was 87% vs. 88% for RB and RH, respectively. RH is not an ideal platform for future multi-centre transplant trials in MCL. RB achieved a 2-year PFS of 81% and a 78% MRD negative rate. Premature closure of the study limited the sample size and the precision of PFS estimates and MRD rates. However, RB can achieve a deep remission and could be a platform for future trials in MCL.

Recent advances in diffuse large B-cell lymphomas (DLBCL) have underscored the importance of tumor microenvironment in escaping host anti-tumor responses. One mechanism is loss of major histocompatibility Class II antigens (MHCII) associated with decreased tumor infiltrating T lymphocytes (TIL) and poor survival. Transcription of MHCII is controlled by CIITA which in turn is regulated by histone acetylation. In this study, we hypothesized that HDAC inhibition with belinostat increases MHCII, CIITA expression, TIL and improves patient outcomes. Primary objective was evaluation of toxicity and response. Twenty-two patients were enrolled for the study. Belinostat was well tolerated with mild toxicity. Two partial responses were observed at 5, 13 months after registration for an overall response rate (ORR) (95% CI) of 10.5% (1.3-33.1%), and three patients had stable disease for 4.7, 42.3+, and 68.4 + months with minimum 3-year follow-up. Included correlative studies support the hypothesis and serve as the basis for SWOG S0806 combining vorinostat with R-CHOP.

CD23 is a trans-membrane protein belonging to the C-type lectin family, widely expressed in the immune system. Originally identified as the low-affinity receptor for IgE, CD23 was later shown to also bind CD21, MHCII and various integrins, and to have pleiotropic roles, including modulation of cell proliferation, differentiation, Ig and cytokine secretion. Expression of membrane CD23 (mCD23) is regulated at least in part by its shedding from the cell surface via activity of metalloproteinases, the most important of which is ADAM10. Interestingly, secreted CD23 (sCD23) isoforms can exert immunomodulatory functions as soluble factors. B cells upregulate mCD23 and serum sCD23 levels have been found to be elevated in inflammatory and autoimmune conditions, including rheumatoid arthritis, and in B cell chronic lymphocytic leukemia. However, the mechanisms responsible for CD23 modulation on B cells are still unclear. Here we show that mCD23 is upregulated on mouse and human B cells found in inflamed lymphoid tissues. Furthermore, in vitro experiments demonstrate that B cell mCD23 upregulation is strictly dependent on soluble signals provided by stromal cells from inflamed, but not non-inflamed lymph nodes, and can be reversed by ADAM10-specific inhibitors. We propose therefore that the balance between B cell mCD23 and sCD23 expression in inflammatory conditions is modulated by microenvironmental signals provided by stromal cells within lymphoid tissues.

A unique population of CD23+ CD21high B cells in inflamed nodes (Bin) has been shown to accumulate in lymph nodes (LNs) draining inflamed joints of TNF‐transgenic (TNF‐tg) mice. Bin cells contribute to arthritis flare in mice by distorting node architecture and hampering lymphatic flow, but their existence in human inflamed LNs has not yet been described. Here, we report the characterization of resident B‐cell populations in fresh popliteal lymph nodes (PLNs) from patients with severe lower limb diseases (non‐RA) and rheumatoid arthritis (RA) patients, and from banked, cryopreserved reactive and normal human LN single cell suspension samples. Bin‐like B cells were shown to be significantly increased in reactive LNs, and strikingly elevated (>30% of total) in RA samples. Histopathology and immunofluorescence analyses were consistent with B follicular hyperplasia and histological alterations in RA vs. non‐RA PLNs. This is the first description of Bin‐like B cells in human inflamed LNs. Consistent with published mouse data, this population appears to be associated with inflammatory arthritis and distortion of LN architecture. Further analyses are necessary to assess the role of CD23+CD21hi Bin‐like B cells in RA pathogenesis and arthritic flare.

Bortezomib is active in mantle cell lymphoma (MCL), with approval in upfront and relapsed settings. Given inevitable recurrence following induction chemoimmunotherapy, maintenance approaches are a rational strategy to improve clinical outcomes. We conducted a phase II study to evaluate the safety and efficacy of six cycles of R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone) plus bortezomib (1.3 mg/m2 days 1 and 4 of 21 d cycles) followed by bortezomib maintenance (1.3 mg/m2 days 1, 4, 8, and 11 every 3 months for 2 years). Sixty-five eligible patients were enrolled. The treatment was well tolerated and toxicities were mainly haematological. The rate of grade ≥3 peripheral neuropathy was low (5%). With a median follow-up of 6.8 years, 2-year progression-free survival (PFS) was 62%, and 2-year overall survival (OS) was 85%. At 5 years, PFS was 28% and OS was 66%. MCL International Prognostic Index scores were significantly associated with 2-year PFS, but did not predict long-term (≥5-year) PFS. Baseline Ki-67 index was significantly associated with survival. Combination R-CHOP with bortezomib followed by maintenance bortezomib appears to improve outcomes compared historically with R-CHOP alone, with prolonged remissions in a subset of patients. These results suggest that inclusion of bortezomib with induction chemotherapy and/or maintenance is promising in MCL and warrants further exploration.

Abstract Introduction: Aggressive induction chemotherapy followed by autologous stem cell transplant (ASCT) is effective for younger patients with mantle cell lymphoma (MCL). Among patients undergoing ASCT, cytarabine-based induction regimen induced higher rates of MRD-negativity compared to R-CHOP (Hermine et al, ASH 2012), and achievement of MRD-negative status was predictive of good outcome. R-bendamustine (RB) has superior efficacy compared to R-CHOP (Rummel et al, Lancet 2013; Flinn et al, Blood 2014) but, there are no data regarding its impact on MRD. S1106 (SWOG) was conducted to compare R-HyperCVAD/MTX/ARAC (RH) or RB to identify the better induction regimen followed by ASCT. We report the result of MRD analysis and updated 2 year PFS/OS. Method: S1106 was a US intergroup (SWOG/ECOG/CALGB), randomized phase II trial. The primary objective was to estimate 2 year PFS, with secondary objectives to assess response rates, OS, toxicities, and MRD status. Inclusion criteria were: untreated stage III, IV or bulky stage II MCL, Cyclin D1 +, age > 18 < 65, and adequate organ function. Randomization was stratified by MIPI. Patients received either 4 cycles of RH or 6 cycles of RB, followed by ASCT. MRD was assessed at baseline and post induction. Genomic tumor DNA was extracted from FFPE tissue or bone marrow aspirate mononuclear cells. PCR amplification of IGH-VDJ, IGH-DJ, and IGK regions was performed followed by high-throughput sequencing to determine the tumor clonotype(s) (Adaptive Biotechnologies). DNA from peripheral blood mononuclear cells (PBMC) and plasma was amplified and sequenced to determine lymphoma molecules per million diploid genomes. Results: A total of 53 patients were accrued out of planned 160. This study was closed prematurely based on predetermined criteria of stem cell mobilization failures on the RH arm. Table 1 shows patient characteristics. The ORR was 94% in RH vs. 83% in RB. The CR rates were 35% (RH) and 40% (RB). Only 4/17 patients on RH and 21/35 patients in RB underwent ASCT; the rest could not complete study (Table 1). RH induced significantly more grade 3/4 heme toxicities as compared to RB (Table 1). The median follow up was 29 months in RH and 26 months in RB. The estimated 2 year PFS was 81% (Fig 1) and OS was 87% for both arms. 27 patients consented to the optional MRD assessment, with 12 paired serial samples (baseline and post induction). 10 were in RB and 2 were in RH. Both patients in RH were MRD positive at baseline and achieved MRD negative status. 1/10 patient in RB was MRD negative at baseline and remained negative throughout treatment. 8/9 patients in RB with baseline MRD-positivity converted to MRD-negative status by the end of induction, and 3/8 did not go to ASCT. Three additional patients were missing baseline samples but were MRD negative at the end of RB. Overall, the estimated 2 year PFS was 100% for all 13 patients who achieved MRD negative status at the end of RB. Conclusions: RH is not an ideal platform for future transplant trials in MCL due to stem cell mobilization failures. RB achieved a 2 year PFS of 81%, higher than the planned target of 75%. It also achieved an 89% MRD negative rate on all the paired samples tested. Low CR on RB could be due to lack of mandatory PET. All patients with MRD negative status remain in remission, with some not having undergone ASCT. Premature closure of the study limited the sample size and the precision of PFS estimates and MRD. However, this analysis suggests that RB can achieve a deep remission and could be a platform for future trials in MCL. Table 1. RH (n=17) RB (n=35) Age 59 (44-66) 57 (33-64) Male (p=0.003) Female 9 8 32 3 Performance status 0 1 11 6 26 9 Disease Stage III IV 1 16 3 32 B sx Yes No 6 11 10 25 BM involvement Positive Negative 14 3 31 4 Extranodal involvement Yes No 15 2 32 3 KI 67 <10% 10-30% >30% 20% 60% 20% 14% 66% 20% MIPI Score Intermediate/High Risk Low Risk 6 11 13 22 Grade 3/4 Hematological toxicities (Induction only) Anemia 56% Neutropenia 63% Febrile neutropenia 31% Thrombocytopenia 69% Anemia 8.6% Neutropenia 34% Febrile neutropenia 14% Thrombocytopenia 17% Reasons for early off treatment or not going to ASCT Failure to collect stem cell 5 Thrombocytopenia 4 Pancytopenia 1 Others 2 Patient choice 4 Progressive disease 2 Failure to collect stem cell 1 Allergy 1 Seizure 1 Insurance denial 1 Others 3 Figure 1. Figure 1. Disclosures Chen: genentech: Consultancy, Speakers Bureau; Seattle Genetics: Consultancy, Research Funding, Speakers Bureau; Merck: Consultancy, Research Funding; Millennium: Consultancy, Research Funding, Speakers Bureau. Forman:Mustang Therapeutics: Research Funding. Cashen:Celgene: Speakers Bureau. Blum:cephalon: Research Funding; Pharmacyclics: Research Funding; Janssen: Research Funding; Celgene: Research Funding. Fenske:Pharmacyclics: Honoraria; Millennium/Takeda: Research Funding; Seattle Genetics: Honoraria; Celgene: Honoraria. Cheson:Celgene: Consultancy, Research Funding; Spectrum: Consultancy; Astellas: Consultancy; Gilead: Consultancy, Research Funding; Roche/Genentech: Consultancy, Research Funding; MedImmune: Research Funding; Ascenta: Research Funding; Pharmacyclics: Consultancy, Research Funding; AstraZeneca: Consultancy; Teva: Research Funding. Smith:Pharmacyclics: Consultancy; Celgene: Consultancy. Faham:adaptive biotech: Employment, Other: stockholders. Wilkins:adaptive biotech: Employment. Leonard:teva: Consultancy; genentech: Consultancy. Kahl:Genentech: Consultancy; AbbVie: Research Funding; Teva: Consultancy.

B-cell malignancies are a common type of cancer. One approach to cancer therapy is to either increase oxidative stress or inhibit the stress response systems on which cancer cells rely. In this study, we combined nontoxic concentrations of Auranofin (AUR), an inhibitor of the thioredoxin system, with nontoxic concentrations of buthionine-sulfoximine (BSO), a compound that reduces intracellular glutathione levels, and investigated the effect of this drug combination on multiple pathways critical for malignant B-cell survival. Auranofin interacted synergistically with BSO at low concentrations to trigger death in multiple malignant B-cell lines and primary mantle-cell lymphoma cells. Additionally, there was less toxicity toward normal B cells. Low AUR concentrations inhibited thioredoxin reductase (TrxR) activity, an effect significantly increased by BSO cotreatment. Overexpression of TrxR partially reversed AUR+BSO toxicity. Interestingly, the combination of AUR+BSO inhibited nuclear factor kappa B (NF-kappa B) signaling. Moreover, synergistic cell death induced by this regimen was attenuated in cells overexpressing NF-kappa B proteins, arguing for a functional role for NF-kappa B inhibition in AUR+BSO-mediated cell death. Together, these findings suggest that AUR+BSO synergistically induces malignant B-cell death, a process mediated by dual inhibition of TrxR and NF-kappa B, and such an approach warrants further investigation in B-cell malignancies. Copyright (C) 2015 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc.

In searching for small-molecule compounds that inhibit proliferation and survival of diffuse large B-cell lymphoma (DLBCL) cells and may, therefore, be exploited as potential therapeutic agents for this disease, we identified the commonly used and well-tolerated antibiotic doxycycline as a strong candidate. Here, we demonstrate that doxycycline inhibits the growth of DLBCL cells both in vitro and in mouse xenograft models. In addition, we show that doxycycline accumulates in DLBCL cells to high concentrations and affects multiple signaling pathways that are crucial for lymphomagenesis. Our data reveal the deneddylating activity of COP-9 signalosome (CSN) as a novel target of doxycycline and suggest that doxycycline may exert its effects in DLBCL cells in part through a CSN5-HSP90 pathway. Consistently, knockdown of CSN5 exhibited similar effects as doxycycline treatment on DLBCL cell survival and HSP90 chaperone function. In addition to DLBCL cells, doxycycline inhibited growth of several other types of non-Hodgkin lymphoma cells in vitro. Together, our results suggest that doxycycline may represent a promising therapeutic agent for DLBCL and other non-Hodgkin lymphomas subtypes.

The biology of follicular lymphoma (FL) is largely dictated by the immune-effector and stromal cells that comprise its tumor microenvironment. FL-infiltrating T-cell populations that are thought to be fundamental to FL biology are follicular helper T-cells (TFH), follicular regulatory T-cells (TFR), a recently described population that regulates TFH activity, and regulatory T-cells (Treg). These T-cell populations have dynamic interactions with mesenchymal stromal cells (MSCs) in the tumor microenvironment. Whereas MSCs have been shown to support FL B-cell and Treg viability, their effects on FL-infiltrating TFH and TFR cells have not been described. Herein we show that MSCs support the viability of FL-infiltrating TFH and TFR, as well as Tregs, in part through an IL-6-dependent mechanism. We further demonstrate that MSCs mediate TFH to TFR conversion by inducing the expression of FoxP3 in TFH cells, demonstrating for the first time that human TFR can be derived from TFH cells. Given that the balance of TFH and TFR populations likely dictate, in part, the biology of this disease, our data support the potential for targeting MSCs as a therapeutic strategy.