Common B-cell non-Hodgkin Lymphomas Research Articles

Research Progress of CAR-T Cell Immunotherapy in B-Cell Non-Hodgkin's Lymphoma--Review

Chimeric antigen receptor T (CAR-T) cell therapy is a rapidly developing new immunotherapy in recent years. Compared with other therapies, CAR-T has significant advantages for high-risk and relapsed/refractory B cell non-Hodgkin's lymphoma (B-NHL) patients. Currently, a variety of anti-CD19 CAR-T cells have been approved by the FDA for the treatment of B-NHL, such as axicabtagene ciloleucel, tisagenlecucel, lisocababtagene maraleucel and brexucabtagene autoleucel. In addition, many studies are actively exploring and developing different targeted CAR-T cells, which show great potential in B-NHL. This review briefly summarized the latest research progress on the application of CAR-T in common B-NHL.

Validation of single nucleotide polymorphisms potentially related to R-CHOP resistance in diffuse large B-cell lymphoma patients.

Aim: Diffuse large B-cell lymphoma (DLBCL) is the most common B-cell non-Hodgkin lymphoma (NHL). Despite the availability of clinical and molecular algorithms applied for the prediction of prognosis, in up to 30%-40% of patients, intrinsic or acquired drug resistance occurs. Constitutional genetics may help to predict R-CHOP resistance. This study aimed to validate previously identified single nucleotide polymorphisms (SNPs) in the literature as potential predictors of R-CHOP resistance in DLBCL patients, SNPs. Methods: Twenty SNPs, involved in R-CHOP pharmacokinetics/pharmacodynamics or other pathobiological processes, were investigated in 185 stage I-IV DLBCL patients included in a multi-institution pharmacogenetic study to validate their previously identified correlations with resistance to R-CHOP. Results: Correlations between rs2010963 (VEGFA gene) and sex (P = 0.046), and rs1625895 (TP53 gene) and stage (P = 0.003) were shown. After multivariate analyses, a concordant effect (i.e., increased risk of disease progression and death) was observed for rs1883112 (NCF4 gene) and rs1800871 (IL10 gene). When patients were grouped according to the revised International Prognostic Index (R-IPI), both these SNPs further discriminated progression-free survival (PFS) and overall survival (OS) of the R-IPI-1-2 subgroup. Overall, patients harboring the rare allele showed shorter PFS and OS compared with wild-type patients. Conclusions: Two out of the 20 study SNPs were validated. Thus, these results support the role of previously identified rs1883112 and rs1800871 in predicting DLBCL resistance to R-CHOP and highlight their ability to further discriminate the prognosis of R-IPI-1-2 patients. These data point to the need to also focus on host genetics for a more comprehensive assessment of DLBCL patient outcomes in future prospective trials.

Real-World Data on the Clinical Features, Therapy Patterns and Outcomes of 651 Elderly Patients with Diffuse Large B-Cell Lymphoma in Latin America: A Study from the Grupo De Estudio Latinoamericano De Linfoproliferativos (GELL)

Background: Diffuse Large B-Cell Lymphoma (DLBCL) is the most common B-cell non-Hodgkin lymphoma. DLBCL commonly affects patients with comorbidities and the elderly. To date, chemoimmunotherapy remains the standard of care for these patients. Data on patients aged ≥65 years managed in Latin America (LATAM) are scarce. Herein, we examine the clinical, treatment and outcome patterns in older patients with DLBCL, with a focus on the effect of treatment in survival outcomes. Methods: We retrospectively analyzed patients aged ≥65 years with newly diagnosed DLBCL managed at different academic institutions in 8 LATAM countries. Demographic characteristics are reported using descriptive statistics. All patients received treatment; those who completed 6 cycles and underwent either CT or PETCT at the end of therapy were considered evaluable for response. Survival curves were estimated using the Kaplan-Meier method. Logistic and Cox proportional-hazard regression models were used to evaluate parameters associated with response and survival. Results: A total of 651 patients were identified and had sufficient data for analysis; median age was 74.1 years (65-96), 335 (51.5%) were female, 400 (61.4%) had ECOG ≤1, and 426 (65.4%) had advanced disease (stage III-IV). No difference in clinical features and laboratory parameters were identified among patients aged <75 vs ≥75 years (Table). The most common first-line chemoimmunotherapy regimens used were standard dose R-CHOP (n=421, 64.7%), R-CVP (n= 74, 11.4 %), R-mini-CHOP (n= 61, 9.4%), and CHOP (n= 29, 4.5%). Eight patients (1.2%) received R-EPOCH, and 43 (6.6%) best supportive care including some receiving dexamethasone and/or rituximab monotherapy. In all patients, the overall response rate was 64.4% (complete 55.3%, partial 9.1%). Responses could not be evaluated in 219 cases (33.6%) either because of early death secondary to toxicity or lymphoma progression during therapy (n=212, 32.5%), or as a complication of patients' comorbidities (n=16, 2.5%). With a median follow up of 60 months (range 1-100), the median overall survival (OS) time was 56 months (95% CI: 38.7-73.2) with a 5-year OS rate of 50%. The causes of death were lymphoma (n=180, 27.6%), infection (n=72,11.1%), non-infectious toxicity (n=6, 0.9%), ischemic cardiopathy (n=7, 1.1%), heart failure (n=6, 0.9%), cerebrovascular event (n=6, 0.9%), and unknown in 41 cases (6.3%). Given our prior work on the impact of serum albumin in DLBCL patients, we analyzed serum albumin levels in elderly patients with DLBCL. Hypoalbuminemia (albumin <3.5mg/dL) was associated with a higher frequency of advanced disease (n=211, 37.8% vs n=152, 27.2%; [p=0.005]), high-risk IPI score (n=77, 14.7% vs n=62, 11%; [p=0.000]), and death (n=147, 22.5% vs n=128, 19.6%; [p=0.000]). In the univariate analysis, the factors influencing mortality were: ECOG ≤1 (p=0.000, HR 1.25[95% IC: 1.0-1.4]), hypoalbuminemia (serum albumin <3.5mg/dL; p=0.031, HR 1.24 [95% IC: 1.02 -1.51]), the use of anthracyclines (p=0.002, HR 0.63, [95 % IC: 0.47 -0.84]), and no achieving a complete response (CR) to first-line treatment (p=0.000, HR 5.01 [95 %IC: 4.25 -6.0]. In the multivariate analysis, all factors remained statistically significant: ECOG ≤1 (p=0.000, aHR 1.32[95 IC: 1.1 -1.5]), hypoalbuminemia (p=0.005, aHR 1.33 [95 % IC: 1.08 -1.6]), the use of anthracyclines (p=0.006, aHR 0.66 [95 % IC: 0.49 -0.88]), and no achieving CR to first-line treatment (p=0.000, aHR 4.91 [95 %IC: 4.1 -5.7]. Conclusions: To our knowledge, this is one of the largest real-world studies on elderly patients with DLBCL in LATAM. In this cohort, survival rates were comparable to those previously reported in the literature. This statement was also true when evaluating the response rates to first-line therapy. We found that ECOG performance status, serum albumin levels, the use of anthracycline and response to first-line therapy were all independently associated to survival. Although the main limitation of this study is its retrospective design, we report that current practices in LATAM remain appropriate for our context and known limitations, thus, this study provides some evidence to clinicians on the therapy patterns and outcomes of such therapies in elderly patients with DLBCL living in resource-limited settings.

Prevalence of IRF4 rearrangement in large B-cell lymphomas of the Waldeyer’s ring in adults

Diffuse large B-cell lymphoma (DLBCL) is the most common B-cell non-Hodgkin lymphoma (B-NHL) in adults. These lymphomas are classified according to gene expression profiling (GEP) into germinal center B-cell (GCB) and activated B-cell type (ABC). Recent studies have suggested new subtypes of large B-cell lymphoma, based on genetic and molecular alterations, among them is large B-cell lymphoma with IRF4-rearrangement (LBCL-IRF4). We used fluorescence in situ hybridization (FISH), GEP (using the DLBCL COO assay by HTG Molecular Inc), and next generation sequencing (NGS) to comprehensively characterize 30 cases of LBCLs located in Waldeyer’s ring in adult patients and to identify LBCL-IRF4. FISH revealed breaks of IRF4 in 2/30 cases (6.7%), BCL2 breaks in 6/30 cases (20.0%), and IGH breaks in 13/29 cases (44.8%). GEP classified 14 cases each as GCB or ABC subtype, and 2 cases remained unclassified; this was concordant with the immunohistochemistry (IHC) in 25/30 cases (83.3%). A subgrouping, based on GEP, was performed: group 1 included 14 GCB cases with the most frequent mutations in BCL2 and EZH2 in 6/14 cases (42.8%). The two cases with IRF4 rearrangement were assigned to this group by GEP and showed IRF4 mutations, supporting the diagnosis of LBCL-IRF4. Group 2 included 14 ABC cases; the most frequent mutations were CD79B and MYD88 identified in 5/14 patients (35.7%). Group 3 included 2 unclassifiable cases in which no molecular patterns were detected. Overall, LBCLs of Waldeyer’s ring in adult patients are a heterogeneous group, including LBCL-IRF4, which shares several features with cases in the pediatric population.

Impact of Circulating Lymphoma at Diagnosis on Outcomes in Patients with Diffuse Large B-Cell Lymphoma

Introduction Diffuse large B-cell lymphoma (DLBCL) is the most common B-cell non-Hodgkin lymphoma. The risk stratification tools continue to evolve and currently, there are various known prognostic factors including revised international prognostic index (R-IPI), MYC/BCL2 overexpression by IHC, MYC/BCL2, and or BCL6 rearrangement (double hit/triple hit lymphomas [DHL/THL]), CD5 expression, and metabolic tumor volume. Outside of case reports and small case series (Muringampurath-John, D et al BJH 2012), the relevance of circulating lymphoma (CL) in patients with DLBCL at diagnosis is largely unknown. Hence, we sought to evaluate the impact of CL at diagnosis on outcomes in patients with DLBCL. Methods This is a single-center, retrospective cohort study of DLBCL patients treated at Ohio State University and included patients who were ≥18 years old, diagnosed with DLBCL from 2010-2021, and had information on flow cytometry at diagnosis. Patients were grouped according to positive or negative flow cytometry results as CL+ and CL-, respectively. The primary endpoint was progression-free survival (PFS), and the secondary endpoint was overall survival (OS) in CL+ and CL- groups. PFS was defined as the time from the start of first-line therapy until lymphoma relapse/progression or death from any cause, censoring at the last clinical assessment. OS was defined as the time from the start of first-line therapy until death or last follow-up. The first-line therapy was categorized into two groups, R-CHOP, and intensive induction therapy (R-EPOCH, R-hyperCVAD, and R-CODOX-M/R-IVAC). Results Among the 181 eligible patients with newly diagnosed DLBCL, 30 (17%) had detectable CL at diagnosis, while 151 (83%) did not. Patients in the CL+ group had a higher median age (66 vs 64 years), more advanced stage disease (97% vs 83%), and R-IPI score of 3-5 (89% vs 62%) compared to the CL- group. The two groups were balanced in regard to gender, race, ECOG performance status, bulky disease, presence of B symptoms, cell of origin, synchronous central nervous system (CNS) involvement, DHL/THL, and first-line therapy. The median PFS was 4.2 years (95%CI=0.9-not reached [NR]) in the CL+ group compared to 10.9 years (95%CI=4.8-NR) in the CL- group (p=0.20, Figure 1). While the median PFS was not significantly different between CL+ and CL- groups among patients treated with R-CHOP chemotherapy (median PFS NR vs 10.9 years, respectively, p=0.99). However, compared to the CL+ group, the median PFS was significantly longer in the CL- group receiving intensive induction therapy (median PFS 1.8 years vs NR, respectively, p=0.04). After adjusting for factors associated with PFS in multivariable cox regression (gender, R-IPI, first-line therapy, synchronous CNS involvement, and DHL/THL), there was no difference in PFS between CL+ and CL- groups (HR=1.48, 95%CI=0.77-2.85, p=0.24). The median OS was 4.2 years (95%CI=1.7-NR) in the CL+ group while it was NR in the CL- group (p=0.05, Figure 2). After adjusting for factors associated with PFS in multivariable cox regression (gender, R-IPI, first-line therapy, synchronous CNS involvement, and DHL/THL), there was no difference in OS between CL+ and CL- groups (HR=1.85, 95%CI=0.90-3.79, p=0.09). Conclusions This is the first study to compare the outcomes of patients with CL+ and CL- groups in newly diagnosed DLBCL. In this largest study to date evaluating the prognostic relevance of CL in patients with DLBCL, we found that CL at diagnosis was not associated with a shorter PFS but trended towards inferior OS. The CL+ group represents a potentially high-risk patient population and needs to be prioritized for experimental and cellular therapies at first relapse. Future prospective studies should include testing for CL and validate the findings found in our study. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

A comprehensive retrospective cohort study of the journey of B-cell lymphoma in Taiwan

Complete disease journey and risk factors for poor outcomes are needed to facilitate effectiveness evaluations of new therapies and clinical decision-making in B-cell Non-Hodgkin lymphoma (B-NHL), particularly in Asia where such data are lacking. This retrospective cohort study used electronic medical records from a regional medical centre in southern Taiwan to follow-up 441 patients newly diagnosed with common B-NHL subtypes: Diffuse Large B-cell Lymphoma (DLBCL), Follicular Lymphoma (FL), Chronic Lymphocytic Leukaemia/Small Lymphocytic Lymphoma (CLL/SLL), Marginal Zone Lymphoma (MZL), Mantle Cell Lymphoma (MCL), and Waldenström macroglobulinemia (WM), between 01-Jan-2008 and 31-Dec-2013, until 31-Dec-2017. Treatment pathways were modelled using a Markov approach. Stage III/IV disease at diagnosis was frequent for patients with DLBCL, FL, MCL and WM. Hepatitis B surface antigen/hepatitis C virus seropositivity was 18.6%/12.3%. Clinical responses to 1st-line treatment were observed in 76.0% (DLBCL), 87.3% (FL), 86.0% (MZL), 60.0% (MCL), and 42.9% (WM) of patients. For DLBCL, disease control was achieved by ~ 50% after 1st-line, ~ 24% after 2nd-line, ~ 17% after 3rd-line. Patients with Stage III/IV DLBCL or age > 65 years at diagnosis had lower rates of active treatment, poorer disease control and higher mortality than patients with early stage disease or age ≤ 65 years. Disease flare < 6 months after 1st-line treatment was significantly associated with mortality. Despite good clinical response rates for some sub-types, survival remains poor. New treatments are needed to improve the outcome of B-NHL.

Lymphoblastic Transformation of Follicular Lymphoma: A Case Report and Review of the Literature

Abstract Follicular lymphoma (FL) is one of the most common B-cell non-Hodgkin lymphomas in adults in the Western world. Prognosis is correlated with the clinical stage and histologic grade and, increasingly, the tumor genetic profile. A subset of patients with FL experiences histologic disease progression (ie, histologic transformation), of which the majority will demonstrate diffuse large B-cell lymphoma. We describe a patient initially diagnosed with low-grade FL with relapsed disease presenting as acute renal failure due to diffuse abdominal lymphadenopathy. Excisional biopsy of an inguinal lymph node at relapse showed high-grade areas with sheets of immature-appearing lymphoid cells adjacent to nodular areas characteristic of low-grade FL. Cells of both components were positive for BCL2 and CD19. The cells of the high-grade component were positive for CD99 and TdT and negative for CD20, whereas cells of the low-grade component were positive for CD20 and negative for CD99 and TdT. Fluorescence in situ hybridization studies demonstrated the IGH/BCL2 rearrangement in addition to an MYC rearrangement in both low- and high-grade components.

Specific Lipid and Metabolic Profiles of R-CHOP-Resistant Diffuse Large B-Cell Lymphoma Elucidated by Matrix-Assisted Laser Desorption Ionization Mass Spectrometry Imaging and in Vivo Imaging.

Diffuse large B-cell lymphoma (DLBCL) is the most common B-cell non-Hodgkin lymphoma. To treat this aggressive disease, R-CHOP, a combination of immunotherapy (R; rituximab) and chemotherapy (CHOP; cyclophosphamide, doxorubicin, vincristine, and prednisone), remains the most commonly used regimen for newly diagnosed DLBCLs. However, up to one-third of patients ultimately becomes refractory to initial therapy or relapses after treatment, and the high mortality rate highlights the urgent need for novel therapeutic approaches based upon selective molecular targets. In order to understand the molecular mechanisms underlying relapsed DLBCL, we studied differences in the lipid and metabolic composition of nontreated and R-CHOP-resistant tumors, using a combination of in vivo DLBCL xenograft models and mass spectrometry imaging. Together, these techniques provide information regarding analyte composition and molecular distributions of therapy-resistant and sensitive areas. We found specific lipid and metabolic profiles for R-CHOP-resistant tumors, such as a higher presence of phosphatidylinositol and sphingomyelin fragments. In addition, we investigated intratumor heterogeneity and identified specific lipid markers of viable and necrotic areas. Furthermore, we could monitor metabolic changes and found reduced adenosine triphosphate and increased adenosine monophosphate in the R-CHOP-resistant tumors. This work highlights the power of combining in vivo imaging and MSI to track molecular signatures in DLBCL, which has potential application for other diseases.

Multiplex Droplet Digital PCR Quantification of Recurrent Somatic Mutations in Diffuse Large B-Cell and Follicular Lymphoma.

A plethora of options to detect mutations in tumor-derived DNA currently exist but each suffers limitations in analytical sensitivity, cost, or scalability. Droplet digital PCR (ddPCR) is an appealing technology for detecting the presence of specific mutations based on a priori knowledge and can be applied to tumor biopsies, including formalin-fixed paraffin embedded (FFPE) tissues. More recently, ddPCR has gained popularity in its utility in quantifying circulating tumor DNA. We have developed a suite of novel ddPCR assays for detecting recurrent mutations that are prevalent in common B-cell non-Hodgkin lymphomas (NHLs), including diffuse large B-cell lymphoma, follicular lymphoma, and lymphoplasmacytic lymphoma. These assays allowed the differentiation and counting of mutant and wild-type molecules using one single hydrolysis probe. We also implemented multiplexing that allowed the simultaneous detection of distinct mutations and an "inverted" ddPCR assay design, based on employing probes matching wild-type alleles, capable of detecting the presence of multiple single nucleotide polymorphisms. The assays successfully detected and quantified somatic mutations commonly affecting enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2) (Y641) and signal transducer and activator of transcription 6 (STAT6) (D419) hotspots in fresh tumor, FFPE, and liquid biopsies. The "inverted" ddPCR approach effectively reported any single nucleotide variant affecting either of these 2 hotspots as well. Finally, we could effectively multiplex hydrolysis probes targeting 2 additional lymphoma-related hotspots: myeloid differentiation primary response 88 (MYD88; L265P) and cyclin D3 (CCND3; I290R). Our suite of ddPCR assays provides sufficient analytical sensitivity and specificity for either the invasive or noninvasive detection of multiple recurrent somatic mutations in B-cell NHLs.

The Synergistic Effect of Venetoclax Combined with a CDK9 Inhibitor in Primary Diffuse Large B Cell Lymphoma and Follicular Lymphoma Cells

The early phase studies have shown the high response rates in chronic lymphocytic leukemia (CLL) patients treated with the BH3 mimetic venetoclax (ABT-199). It indicated that inhibition of BCL-2 is a viable strategy for the treatment of lymphoid malignancies. Objective anti-tumor responses were also observed in patients with other common B-cell non-Hodgkin lymphomas (NHLs) such as follicular lymphoma (FL) or diffuse large B-cell lymphoma (DLBCL), however the overall response rates are not as high as those in CLL patients. Targeting only one anti-apoptotic protein may lead to or uncover resistance owing to activity of other anti-apoptotic BCL2-family members in these settings. MCL-1 is associated with both intrinsic and acquired resistance to venetoclax and thus inhibition of MCL-1 is being explored through either direct inhibition or indirect targeting. Expression of MCL-1 is maintained via p-TEFb-mediated transcription, of which CDK9 plays a critical role. Here we aimed to investigate the combined effects of CDK9 inhibitor and venetoclax in primary DLBCL and FL cells.Inhibition of CDK9 via a small molecule A-1467729.0 (AbbVie) caused rapid loss in phosphorylation (Serine 2) of RNA polymerase II and MCL-1 expression in all tested primary cells of DLBCL and FL patients, confirming the intended effect of CDK9 inhibition. Primary samples from 12 NHL cases (6 DLBCL including 3 GCB/3 non-GCB and 6 FL) were tested for their ex vivo response to A-1467729.0 or venetoclax alone or in combination. Apoptosis assays showed negligible effects (<10% induction) of A-1467729.0 at concentrations of 1, 10, 50 and 100 nM in 11 of 12 samples, while venetoclax at same dose range induced dose-dependent apoptosis in all samples. At 10 nM, the range of venetoclax-induced apoptosis was 16.8% - 55.3% (median 27.9%). Co-treatment with venetoclax and A-1467729.0 demonstrated synergistic effects at multiple doses in all 12 samples as evidenced by flow cytometry based apoptotic assay and PARP cleavage. Quantitative flow cytometry (QFC) studies (molecules/cell) showed that DLBCL and FL patient samples had comparable levels of anti-apoptotic proteins including BCL-2, MCL-1 and BCL-xl. However, BIM levels were higher in DLBCL in comparison to FL samples. Immunohistochemical staining of BIM in formalin-fixed paraffin embedded tissues confirmed this trend. Interestingly, venetoclax was more potent in inducing apoptosis in DLBCL patient samples than FL patient samples ex vivo. QFC data revealed a correlation between 1) BCL-2/BIM ratio and IC50 of venetoclax; 2) BCL-2/(MCL-1+BCL-xl) ratio and IC50 of venetoclax in FL. Importantly, venetoclax and CDK9 inhibitor combination demonstrated superior anti-tumor efficacy in xenograft mouse model of B-cell NHL than either agent alone.In summary, small molecule inhibition of CDK9 in primary NHL cells results in rapid down regulation of MCL1 expression. A-1467729.0, in combination with venetoclax, demonstrates synergistic activity as shown by apoptosis induction in primary DLBCL and FL cells. QFC of BCL-2 family proteins may be a useful biomarker for predicting response to BCL-2 inhibition in FL. The in vitro synergy between CDK9 inhibitor and venetoclax was also seen in vivo xenograft studies. These data support further investigation for combination therapy with CDK9 inhibitor and venetoclax in B-cell NHL. DisclosuresPhillips:AbbVie: Employment, Other: Shareholder, Patents & Royalties. Souers:AbbVie: Employment, Equity Ownership. Tapang:AbbVie: Employment, Equity Ownership. Albert:AbbVie: Employment, Equity Ownership.

Clinical significance of acquired cytogenetic clones in patients with treated follicular lymphoma.

Follicular lymphoma (FL) is the second most common B-cell non-Hodgkin lymphoma worldwide. In most patients, the disease is diagnosed at advanced stages and cannot be cured using conventional therapeutic approaches. To assess the role of cytogenetic abnormalities in therapy-related myeloid neoplasms (tMNs), we studied the clinicopathologic and cytogenetic features of treated FL patients who subsequently developed a new acquired cytogenetic clone (ACC). Twenty-five treated FL patients developed new cytogenetic abnormalities from 2009 to 2012. Patients were divided into 3 groups based on the presence and absence of tMNs: group 1, ACC without tMNs after a median follow-up of 15 months; group 2, ACC with possible tMN after silent ACC detection; group 3, tMNs present at the first ACC detection. The most frequent cytogenetic aberrations involved chromosome 7. Compared with group 1, group 3 had significantly greater size of ACC, higher frequency of chromosome 7 aberrations, more likely showed dysplasia, and lower platelet count (P = .03). Our results indicate that the presence of ACC alone is insufficient for diagnosis of tMNs. The proportion of cells with specific aberrations at first ACC, bone marrow dysplasia, and low platelet counts might predict outcome of ACC.

MicroRNA dysregulation in B-cell non-Hodgkin lymphoma

MicroRNA dysregulation in B-cell non-Hodgkin lymphoma Emilia L Lim,1 Marco A Marra1,21Canada&rsquo;s Michael Smith Genome Sciences Centre, BC Cancer Agency, Vancouver, BC, Canada; 2Department of Medical Genetics, University of British Columbia, Vancouver, BC, CanadaAbstract: B-cell non-Hodgkin lymphomas (NHLs) are lymphoproliferative disorders that can arise at different stages of B-cell development. Even though molecular classification of NHL has allowed for more accurate recognition of distinct aggressive lymphoma subtypes, many patients still fail to respond to standard therapy. As such, there is a need to identify biomarkers and therapeutic targets that can lead to more specific treatments for each NHL patient&#39;s disease. MicroRNAs (miRNAs) are small, 17&ndash;25 nt RNA molecules that regulate gene expression at the posttranscriptional level. miRNA expression and function is often coordinately dysregulated in NHL, and consequently results in each NHL disease type harboring a distinct miRNA expression signature. miRNA dysregulation may be a consequence of several mechanisms, ranging from dysregulation of the DNA sequences encoding the miRNA to transcriptional regulation of miRNA loci, to dysregulation of the miRNA biogenesis pathway or dysregulation of messenger RNA (mRNA) targets. This coordinated dysregulation of miRNA expression systematically results in the activation of several oncogenic pathways, and consequently the reprogramming of B-cell NHL transcriptomes. The widespread dysregulation of miRNAs suggests that miRNAs may be used as a diagnostic and prognostic tool, and also as actionable drug targets. In this review, we summarize the miRNA profiles of the most common B-cell NHLs, discuss the causes and consequences of miRNA dysregulation, and consider the prospects of miRNA-based biomarkers and therapeutic targets in NHL.Keywords: miRNA, non-Hodgkin lymphoma, dysregulation, therapy

IL10 Receptor a Is a Novel Therapeutic Target That Is Epigenetically Disregulated in Low Grade Lymphomas with Plasmacytic Differentiation.

Abstract 2383Waldenstrom's Macroglobulinemia (WM) is the clinical manifestation of lymphoproliferative disorders characterized by a clonal lymphoplasmacytic proliferation, excessive IgM secretion and elevated serum viscosity.Most common B-cell non-Hodgkin lymphomas associated with WM are Lymphoplasmacytic Lymphomas (LPL) and Marginal Zone Lymphomas (MZL), especially those displaying plasmacytic differentiation (MZL-P). In order to elucidate common pathogenetic mechanisms that result in WM across lymphoma types, we profiled the methylome of 6 LPLs, 10 MZL-Ps, as well as normal B-cell subsets purified from human tonsillar tissue –germinal center B-cell controls (GCB, n=3) and plasma cell (PC, n=3) controls, using HELP assay and high-density oligonucleotide microarray from RocheNImblegen that queries DNA methylation level of 50,000 cytosine residues distributed among 14,000 gene promoters.Unsupervised hierarchical clustering approach using Ward’s method and Eucledian distances separated normal GCB and PC from the lymphoma cases and identified bi-directional changes in gene methylation: with aberrant gain or loss of methylation at specific genomic locations. We further utilized a t-test and identified 208 probesets that were differentially methylated between controls and lymphomas at p<0.01 and mean log-ratio difference between 2 groups >1.5 corresponding to 30% methylation difference. Ingenuity pathway analysis revealed CDKN1A and TGFb networks as the most aberrantly methylated in lymphomas.Remarkably, we observed that Interleukin 10 Receptor a (IL10RA), was aberrantly hypomethylated in both lymphoma subtypes, resulting in its aberrant overexpression. IL10RA is a subunit of the IL10 cytokine receptor that is known to be a key factor in terminating the inflammatory responses via signaling through the JAK/STAT pathway resulting in STAT3 activation, which also plays an integral part in GCB differentiation and commitment to the plasma cell lineage. We thus hypothesized that aberrant epigenetic upregulation of IL10RA might promote survival and expansion of lymphoma cells. We predicted that stimulating IL10RA with IL10 ligand may lead to increased cell growth, while the blockade of IL10RA would inhibit cell growth. We selected 2 anti-IL10RA antibodies, which have previously been reported to have receptor blocking properties in vivo in mice. We tested 1ug/ml, 5ug/ml and 10ug/ml of each antibody and 10 ng/ml of the stimulatory IL10 ligand and observed that ligand provided stimulatory effect on the growth rate of a panel of B-cell lymphoma, including LPL cell line, while both anti-IL10RA antibodies had marked growth inhibitory effects. We further determined that growth inhibition resulted from dramatic induction of apoptosis in cells treated with the blocking anti-IL10RA antibodies. Further studies revealed that induction of apoptosis followed specific inhibition of signaling through JAK1/2 and phosphorylation of STAT3Y705 immediately after treatment and inhibition of signaling through MAPK and phosphorylation of STAT3S727 at later treatment time points.In conclusion, we determined that IL10RA is aberrantly methylated and overexpressed in subtypes of low grade lymphomas exhibiting plasmacytic differentiation and manifesting as WM. Hence, strategies targeting this pathway should be explored as potential therapy for WM (irrespective of the underlying type of lymphoma). Disclosures:No relevant conflicts of interest to declare.

Diagnosis of B-Cell Non-Hodgkin Lymphomas with Small-/Intermediate-Sized Cells in Cytopathology

Fine needle sampling is a fast, safe, and potentially cost-effective method of obtaining tissue for cytomorphologic assessment aimed at both initial triage and, in some cases, complete diagnosis of patients that present clinically with lymphadenopathy. The cytologic diagnosis of B-cell non-Hodgkin lymphomas composed of small-/intermediate-sized cells, however, has been seen as an area of great difficulty even for experienced observers due to the morphologic overlap between lymphoma and reactive lymphadenopathies as well as between the lymphoma entities themselves. Although ancillary testing has improved diagnostic accuracy, the results from these tests must be interpreted within the morphological and clinical context to avoid misinterpretation. Importantly, the recognition of specific cytologic features is crucial in guiding the appropriate selection of ancillary tests which will either confirm or refute a tentative diagnosis. For these reasons, we here review the cytologic characteristics particular to five common B-cell non-Hodgkin lymphomas which typically cause the most diagnostic confusion based on cytological assessment alone: marginal zone lymphoma, follicular lymphoma, mantle cell lymphoma, chronic lymphocytic leukemia/small lymphocytic lymphoma, and lymphoplasmacytic lymphoma. We summarize the most pertinent cytomorphologic features for each entity as well as for reactive lymphoid hyperplasia, contrast them with each other to facilitate their recognition, and highlight common diagnostic pitfalls.