Activated B-cell Subtype Research Articles

Single and Combinatorial Effects of Metformin and Thymoquinone in Diffuse Large B Cell Lymphoma Cells.

Diffuse large B cell lymphoma (DLBCL) is classified into Germinal Center B-cell (GCB) and activated B-cell (ABC) subgroups originating from different stages of lymphoid differentiation. Cell of origin dictates the behavior and therapeutic response of DLBCL. This study aimed to evaluate single and combinatorial effects of metformin and thymoquinone (TQ) in two DLBCL cell lines belonging to GCB and ABC subtypes. Metformin and TQ caused dose-dependent responses in both ABC and GCB DLBCL subtypes. Metformin had a greater impact on the ABC subtype while TQ demonstrated more pronounced effects on the GCB subtype. Synergistic effects were observed in the DHL4 (GCB subtype) but not in the HBL1 (ABC subtype) cell line. This is the first study to compare the effects of metformin and TQ in ABC versus GCB subtype of DLBCL. It brings valuable results that could be utilized in further research aimed at reshaping treatments for subtype-specific lymphomas.

Biomarker potential of nuclear Nrf2 activation in the ABC subtype of diffuse large B‑cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive B-cell lymphoma characterized by distinct subtypes and heterogeneous treatment outcomes. Oxidative stress and the dysregulation of related regulatory genes are prevalent in DLBCL, prompting an investigation into the nuclear factor erythroid 2-related factor 2 (Nrf2)-kelch-like ECH-associated protein 1 (Keap1) signaling pathway and associated genes. The present study assessed pathological specimens and clinical data from 43 newly diagnosed patients with DLBCL, comparing the associations and correlations between the expression of Nrf2, Keap1, microtubule-associated protein 1 light chain 3β (LC3B) and nitrotyrosine and the activated B-cell (ABC) and germinal center B-cell (GCB) subtypes of DLBCL using immunohistochemistry and digital image analysis software. Nuclear Nrf2 activation was observed in 33.3% of patients with DLBCL ABC, demonstrating a higher prevalence of hepatitis B surface antigen positivity, calcium ions and significant body weight loss (P<0.05). Total Nrf2 expression was associated with the DLBCL GCB subtype and inversely correlated with Keap1 expression in the DLBCL ABC subtype. Furthermore, a positive correlation was demonstrated between Nrf2 and LC3, indicating that total Nrf2 is inhibited by Keap1 and regulates LC3 expression. The ABC subtype was also associated with lower white blood cell counts and more frequent chemotherapy courses than the GCB subtype. These findings suggest that nuclear Nrf2 could be a biomarker for DLBCL clinical diagnosis.

Effect of AVM0703 treatment on the limitations of R-CHOP and survival in a model applicable to ABC subtype DLBCL.

e19060 Background: The current treatment regimen for DLBCL consisting of rituximab-cyclophosphamide-doxorubicin-vincristine-prednisone (RCHOP), is challenged by molecular and prognostic characterization of DLBCL. Activated B-cell (ABC) lymphoma has a significantly worse prognosis (5 yr OS rate: 45% - 56%) than germinal center B-cell (GCB) lymphoma (78% - 80%) after RCHOP treatment. Around 40% of patients ultimately relapse and/or develop primary refractory disease, cardiotoxicity and long-term secondary malignancies. Efforts are being made to improve the RCHOP backbone by including dose intensification and by the addition of new agents to increase the efficacy and reduced toxicity of RCHOP (McKelvey EM Cancer 1976; Wilson WH Blood 2002; Molina TJ JCO 2014; Coiffier B Hematology 2016). AVM0703, an immunomodulatory drug, mobilizes bispecific γδ and inv-TCR double-positive novel T and NKT cells. In a previous study employing an aggressive, immune resistant A20 mouse lymphoma model (Bascuas T J Transl Med. 2016), neoadjuvant AVM0703 was additive/synergistic, when cyclophosphamide/fludarabine (CyFlu) was dosed 3 and 24 hr later, with no additional toxicities. In the present study we combined AVM0703 with RCHOP to evaluate the efficacy in A20 mice model. Methods: 10 week old Balb/c mice were injected with 2x106 murine A20 B-cells in the flank and were randomized (1:1 male: female) into 1) Placebo 2) 3 cycles RCHOP 3) 3 cycles AVM0703 (18mg/kg HED) 4) 3 cycles AVM0703+RCHOP, when tumor size reached ~100 mm3. Mice who achieved complete remission (CR) were re-challenged at 21-25 wks later. Pre and post re-challenge, presence of memory cells was analyzed using flow cytometer. All mice were monitored continuously as per the IACUC protocol. Results: Mice treated with the combination of AVM0703+RCHOP achieved superior CR compared to RCHOP alone, which was not curative in any mice. In first phase of the study, 3 out of 11 AVM0703 +RCHOP treated mice showed CR after only 1-cycle. In second phase, 2 of 12 mice survived in CR for &gt;150 days after 3 cycles, whereas no RCHOP mice survived beyond 2 cycles. AVM0703+RCHOP treated mice in long-term CR, had memory cells (CD44hi-Cd62low) pre- and post re-challenge, and no A20 engraftment, suggesting strong immunological memory induced by AVM0703+RCHOP. Conclusions: AVM+RCHOP demonstrated anti-tumor activity in A20 lymphoma mice, with reduced toxicity, compared to RCHOP. Larger preclinical studies are underway to confirm the activity of AVM0703+RCHOP against RCHOP resistant ABC DLBCL, and the potential to reduce total RCHOP exposure to only 3 cycles while maintaining or improving efficacy. [Table: see text]

Sinonasal DLBCL: molecular profiling identifies subtypes with distinctive prognosis and targetable genetic features

AbstractPrimary sinonasal diffuse large B-cell lymphoma (PSDLBCL) is a rare lymphoma with a variable prognosis and a unique relapse/dissemination pattern involving the central nervous system and skin. The underlying molecular mechanisms leading to this heterogeneity and progression pattern remain uncharted, hampering patient-tailored treatment. To investigate associated mechanisms, we analyzed clinical data and used immunohistochemistry, gene-expression profiling, cytogenetics, and next-generation sequencing in a cohort of 117 patients with PSDLBCL. The distribution in cell-of-origin (COO) was 68 (58%) activated B-cell (ABC), 44 (38%) germinal center B-cell (GCB), and 5 (4%) unclassifiable. COO was significantly associated with progression-free survival (PFS) and lymphoma-specific mortality (LSM) in both the overall cohort (5-year PFS: ABC, 43% vs GCB, 73%; LSM: ABC, 45% vs GCB, 14%) and in the subgroup of patients receiving immunochemotherapy (5-year PFS: ABC, 55% vs GCB, 85%; LSM: ABC, 28% vs GCB, 0%). ABC lymphomas were mainly MCD class, showing a high prevalence of MYD88 (74%) and CD79B (35%) mutations compared with GCB lymphomas (MYD88 23%; CD79B 10%) (P < .01). The ABC subtype frequently displayed cMYC/BCL2 coexpression (76% vs 18% GCB; P < .001) and HLA-II loss (48% vs 10% GCB; P < .001). PD-L1 expression and copy-number alterations were rare. All lymphomas were Epstein-Barr virus-negative. Our data suggest molecular profiling as a potent tool for detecting prognostic subgroups in PSDLBCL, exposing links to known relapse/dissemination sites. The ABC subgroup’s MCD genetic features, shared with lymphomas at other nonprofessional lymphoid sites, make them potential candidates for targeted B-cell and toll-like receptor signaling therapy.

Single-cell mass cytometric analysis of peripheral immunity and multiplex plasma marker profiling of non-small cell lung cancer patients receiving PD-1 targeting immune checkpoint inhibitors in comparison with platinum-based chemotherapy.

The effect of platinum-based chemotherapy (Chem.) and second- or multiple- line immune checkpoint PD-1 blocking therapy by Nivolumab or Pembrolizumab (ICI) was assayed in the peripheral blood of non-small cell lung cancer (NSCLC) patients. Flow cytometry was used to detect NSCLC-related antigen binding IgG antibodies. The Luminex MagPix multiplex bead-based cytokine/chemokine detecting system was used to quantitatively measure 17 soluble markers in the plasma samples. Single-cell mass cytometry was applied for the immunophenotyping of peripheral leukocytes. The incubation of patient derived plasma with human NSCLC tumor cell lines, such as A549, H1975, and H1650, detected NSCLC-specific antibodies reaching a maximum of up to 32% reactive IgG-positive NSCLC cells. The following markers were detected in significantly higher concentration in the plasma of Chem. group versus healthy non-smoker and smoker controls: BTLA, CD27, CD28, CD40, CD80, CD86, GITRL, ICOS, LAG-3, PD-1, PD-L1, and TLR-2. The following markers were detected in significantly higher concentration in the plasma of ICI group versus healthy non-smoker and smoker controls: CD27, CD28, CD40, GITRL, LAG-3, PD-1, PD-L1, and TLR-2. We showed the induction of CD69 and IL-2R on CD4+ CD25+ T-cells upon chemotherapy; the exhaustion of one CD8+ T-cell population was detected by the loss of CD127 and a decrease in CD27. CD19+CD20+, CD79B+, or activated B-cell subtypes showed CD69 increase and downregulation of BTLA, CD27, and IL-2R in NSCLC patients following chemotherapy or ICI. Peripheral immunophenotype caused by chemotherapy or PD-1 blocking was shown in the context of advanced NSCLC.

Cell-of-origin effect of polatuzumab vedotin in diffuse large B-cell lymphoma: no ordinary subgroup analysis

AbstractSubgroup analysis from the POLARIX trial of polatuzumab vedotin plus chemotherapy for untreated large B-cell lymphoma suggests greater efficacy among patients with activated B-cell subtype disease. Both preclinical and additional clinical evidence support this interaction between cell-of-origin and polatuzumab efficacy.

CT Based Tumor Radiomics with Machine Learning Classifiers for Molecular Subtyping of Diffuse Large B Cell Lymphoma.

Diffuse large B-cell lymphoma (DLBL) has been classified into two distinct molecular subtypes based on their cell of origin- namely germinal center B-cell-like (GCB) and activated B-cell-subtype (ABC) based on Han's algorithm. These two entities are also prognostically distinct. We assessed CT based tumor radiomic features using machine learning classifiers to distinguish the two molecular subtypes. One hundred one patients were accrued in the study after institutional ethics committee approval. GCB subtype was diagnosed in 59 while 42 patients were ABC subtype. Lesion with maximum SUV was delineated using a semi-automated segmentation tool (3D slicer) and radiomic features were extracted using Pyradiomics. A total of 1030 features were extracted including Shape features, first, second, third, higher order and wavelet-based features. Recursive feature elimination was used for feature selection and optimal 10 features were selected. Five machine learning (ML) tools were used to build a model to sub- classify the two molecular subtypes. (Table -1) Model performance was assessed using Accuracy, Precision, Recall, F1 score and Area under curve of receiver operating characteristic curve. The models were constructed by two methods.1) A 5-fold internal cross validation strategy & 2) splitting the data into training (70%) and validation set (30%). Recursive feature elimination using a random forest algorithm method was used to select 7 radiomic features. These 7 features were then used to build the model using the 5 ML classifiers. Of the five classifiers, Random Forest Classifier (RFC) using a 70:30 training: test strategy was the best performer among all the ML models, with the highest accuracy of 80%, AUC of 0.87, recall(sensitivity), f1score(specificity) & precision (Positive Predictive value; PPV) or precision of 82% respectively. When an internal 5-fold cross validation strategy was used, RFC again performed better than other ML classifiers with an accuracy of 75%, AUC of 0.80, recall of 79%, PPV of 77% and specificity of 79%. The machine-learning based Radiomics features extracted from pre-treatment CT images can provide a simple and non-invasive method for the prediction of DLBL molecular subtypes with favorable predictive accuracy. The Random Forest Classifier was the most accurate in distinguishing GCB subtype from the ABC subtype.

Leukemic presentation and progressive genomic alterations of MCD/C5 diffuse large B-cell lymphoma (DLBCL).

Diffuse large B-cell lymphoma (DLBCL) is a heterogenous group of lymphoid malignancies. Based on gene expression profiling, it has been subdivided into germinal center (GC)-derived and activated B-cell (ABC) types. Advances in molecular methodologies have further refined the subclassification of DLBCL, based on recurrent genetic abnormalities. Here, we describe a distinct case of DLBCL that presented in leukemic form. DNA sequencing targeting 275 genes revealed pathogenically relevant mutations of CD79B, MyD88, TP53, TBL1XR1, and PIM1 genes, indicating that this lymphoma would be best classified as MCD/C5 DLBCL, an ABC subtype. Despite an initial good clinical response to BTK inhibitor ibrutinib, anti-CD20 antibody rituxan, alkylating agent bendamustine, and hematopoietic stem-cell transplant, the lymphoma relapsed, accompanied by morphologic and molecular evidence of disease progression. Specifically, the recurrent tumor developed loss of TP53 heterozygosity (LOH) and additional chromosomal changes central to ABC DLBCL pathogenesis, such as PRDM1 loss. Acquired resistance to ibrutinib and rituxan was indicated by the emergence of BTK and FOXO1 mutations, respectively, as well as apparent activation of alternative cell-activation pathways, through copy-number alterations (CNAs), detected by high-resolution chromosomal microarrays. In vitro, studies of relapsed lymphoma cells confirmed resistance to standard BTK inhibitors but sensitivity to vecabrutinib, a noncovalent inhibitor active against both wild-type as well as mutated BTK. In summary, we provide in-depth molecular characterization of a de novo leukemic DLBCL and discuss mechanisms that may have contributed to the lymphoma establishment, progression, and development of drug resistance.

CCDC50, an essential driver involved in tumorigenesis, is a potential severity marker of diffuse large B cell lymphoma

Diffuse Large B Cell Lymphoma (DLBCL) is the most common form of blood cancer. Among the subtypes, the activated B-cell (ABC) subtype is typically more aggressive and associated with worse outcomes. However, the underlying mechanisms are not fully understood. In this study, we performed microarray analysis to identify potential ABC-DLBCL-associated genes. We employed Kaplan–Meier methods and cox univariate analysis to explore the prognostic value of the identified candidate gene Coiled-coil domain containing 50 (CCDC50). Additionally, we used DLBCL cell lines and mouse models to explore the functions and mechanisms of CCDC50. Finally, we isolated CCDC50-bearing exosomes from clinical patients to study the correlation between these exosomes and disease severity. Our results demonstrated that CCDC50 not only showed significantly positive correlations with ABC subtype, tumor stage and number of extranodal sites, but also suggested poor outcomes in DLBCL patients. We further found that CCDC50 promoted ABC-DLBCL proliferation in vitro and in vivo. Mechanistically, CCDC50 inhibited ubiquitination-mediated c-Myc degradation by stimulating the PI3K/AKT/GSK-3β pathway. Moreover, CCDC50 expression was positively correlated with c-Myc at protein levels in DLBCL patients. Additionally, in two clinical cohorts, the plasma CCDC50-positive exosomes differentiated DLBCL subtypes robustly (AUC > 0.80) and predicted disease severity effectively (p < 0.05). Our findings suggest that CCDC50 likely drives disease progression in ABC-DLBCL patients, and the CCDC50-bearing exosome holds great potential as a non-invasive biomarker for subtype diagnosis and prognosis prediction of DLBCL patients.

Efficacy Analysis of Bruton's Tyrosine Kinase Inhibitor Combined Regimen in the Treatment of Elderly Patients with Relapsed/Refractory Primary Central Nervous System Lymphoma

Background Primary central nervous system lymphoma (PCNSL) is a kind of aggressive non-Hodgkin lymphoma that originates in the brain, spinal cord, eyes or pia mater without evidence of systemic involvement, which accounts for about 3% of central nervous system tumors and has a poor prognosis. Most patients will eventually relapse and the 5-year survival rate is 29.6%. Zanubrutinib is a kind of new generation BTK inhibitor, which can penetrate the blood-brain barrier and be used in the treatment of PCNSL. The study aims to investigate the efficacy and safety of a zanubrutinib-based combination regimen in the treatment of elderly relapsed/refractory(R/R) PCNSL. Methods We retrospectively analyzed elderly patients with R/R PCNSL who received zanubrutinib-based combination regimen at the Affiliated Hospital of Nantong University over the past few years. We collected the information of patients diagnosed with PCNSL in the SEER project of the National Cancer Institute (NCI), and analyze the survival status. We used CCK8 and EdU to detect the effect of zanubrutinib on the viability and proliferation of activated B-cell subtype (ABC) diffuse large B-cell lymphoma (DLBCL) cell lines with genotypes similar to PCNSL. Results The 7 elderly R/R PCNSL patients treated with zanubrutinib-based regimen (zanubrutinib, rituximab and temozolomide) showed an overall response rate (ORR) of 85.7% (6/7), a complete response (CR) of 57.1% (4/7) and a partial response (PR) of 28.6% (2/7), and 14.3% (1/7) of patients experienced disease progression (PD). Patients' gender, KPS score, ECOG score, surgical treatment or not, positive cerebrospinal fluid protein (all P>0.05) were not significantly associated with the treatment effect. Pathological type and IESLG score were related to treatment effect (P<0.05). The median progression-free survival (PFS) and median overall survival (OS) of patients in this study were not reached. Through the analysis of clinical data of PCNSL patients in SEER database from 1975 to 2016, it is concluded that elderly patients (age>60 years old) and male PCNSL patients have a worse prognosis, the primary site of PCNSL, treatment methods and the choice of the disease have impact on the prognosis, and there is no significant difference between the disease and other causes in terms of the cause of death. In the survival analysis of elderly patients, gender, surgery and radiotherapy sequence had no significant effect on prognosis, and patients who received chemotherapy had better OS than those who received radiotherapy. The cell viability of ABC subtype DLBCL cell line OCI-Ly10 decreased in a concentration- and time-dependent manner with zanubrutinib and temozolomide (P<0.05). The inhibitory effect of zanubrutinib and temozolomide on the proliferation of OCI-Ly10 cell line was stronger than that of single drug. Conclusion Zanubrutinib-based combination therapy has good efficacy and controllable safety in elderly R/R PCNSL patients. The prognosis of elderly patients is poor, and the OS of chemotherapy patients is better than that of radiotherapy among the treatment options in the pre-BTKi era. Zanubrutinib inhibits cell viability and cell proliferation in ABC subtype DLBCL cell line OCI-Ly10.

Transformed Waldenström Macroglobulinemia: Update on Diagnosis, Prognosis and Treatment

Histological transformation (HT) to an aggressive lymphoma results from a rare evolution of Waldenström macroglobulinemia (WM). A higher incidence of transformation events has been reported in MYD88 wild-type WM patients. HT in WM can be histologically heterogeneous, although the diffuse large B-cell lymphoma of activated B-cell subtype is the predominant pathologic entity. The pathophysiology of HT is largely unknown. The clinical suspicion of HT is based on physical deterioration and the rapid enlargement of the lymph nodes in WM patients. Most transformed WM patients present with elevated serum lactate dehydrogenase (LDH) and extranodal disease. A histologic confirmation regarding the transformation to a higher-grade lymphoma is mandatory for the diagnosis of HT, and the choice of the biopsy site may be dictated by the findings of the 18fluorodeoxyglucose-positron emission tomography/computed tomography. The prognosis of HT in WM is unfavorable, with a significantly inferior outcome compared to WM patients without HT. A validated prognostic score based on 3 adverse risk factors (elevated LDH, platelet count &lt; 100 × 109/L and any previous treatment for WM) stratifies patients into 3 risk groups. The most common initial treatment used is a chemo-immunotherapy (CIT), such as R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone). The response duration is short and central nervous system relapses are frequent. Whether autologous stem cell transplantation could benefit fit patients responding to CIT remains to be studied.

META-ANALYSIS DISPLAYED TRANSCRIPTOMIC SIGNATURE OF DIFFUSE LARGE B-CELL LYMPHOMA WITH SIGNIFICANT DIFFERENCES BETWEEN SUBTYPES

Lymphomas are cancers that affect the hematological system originated in the lymphatic system with an abnormal proliferation of lymphoid cells. Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin lymphoma in adults. This lymphoma is well characterized by the presence of B-lineage lymphoid cells with immature features with diffuse proliferation in the affected tissue. This disease usually follows an aggressive clinical course, placing patients in immediate need of chemotherapeutic treatment. DLBCL can be further classified into two molecular subtypes, center-germinative B-cell (GCB) and activated B-cell (ABC) type lymphomas. This classification indicates the different stages of maturation of the neoplastic cells, and the different pathological mechanisms associated with them. The present work aimed to compare transcriptome signature of the center-germinative B-cell (GCB) and activated B-cell (ABC). Microarray studies were selected based on predefined criteria and downloaded from the GEO database. Data acquisition, processing, meta-data construction, as well as the differentially expressed genes were performed using R studio packages. Also, significantly enriched gene sets were identified using Gene Set Enrichment Analysis (GSEA). Gene expression and survival analysis from TCGA database were performed using the UALCAN and GEPIA webservers. From a total of 7,854 differentially expressed genes, 465 significantly down-regulated genes and 403 up-regulated genes in GCB subtype when compared to ABC subtype. Among these DEGs, in the GCB versus ABC comparison, the genes with the highest expression level were SPIN3 (FC: 1.12E+20), ZNF791 (FC: 9.43E+19) and TSPAN3 (FC: 9.18E+19), while the significant DEGs with the lowest expression level were KMT2C (FC: -3.09E+20), RIOX2 (FC: -1.49E+20) and ABHD11 (FC: -6.96E+19). Interestingly, only two genes had their expression levels significantly correlated to the DLBCL patient survival, the ABHD11 gene (p-value = 0.018) and MORF4L2 (p-value = 0.029). ABHD11 gene at low-medium level and MORF4L2 gene at high expression level significantly decrease the survival of patients. the meta-data generated from this study suggests that patients with diffuse large B-cell lymphoma pattern molecularly relevant gene expression in aspects related to cell proliferation, cell metabolism, and insensitivity to inhibitory signals to tumor progression as presented by the type of activated B cells. The findings converge the significant modulation of germinal center cells to gene expression regulation processes. Further experiments are still required, the transcriptional and translational activity, as well as cellular activity, appear to be decreased in the GCB type. Our findings, such as the one regarding the ABHD11 gene, which at low levels in GCB type, confers decreased survival rate in lymphoma patients. Therefore, this work brings important information for the understanding of the pathological process of the disease and its subtypes, and such biomarkers may serve as potential candidates for the development of molecular diagnostic methods and new therapeutic strategies for DLBCL subgroups.

Next-Generation Sequencing Highlights of Diffuse Large B-cell Lymphoma in a Tertiary Care Hospital in North India.

Introduction: Next-generation sequencing (NGS) elucidates the diffuse large B-cell lymphoma (DLBCL) genetic characteristics by finding recurrent and novel somatic mutations. This observational study attempted to create an NGS panel with a focus on identifying novel somatic mutations which could have potential clinical and therapeutic implications. This panel was created to look for mutations in 133 genes chosen on basis of a literature review and it was used to sequence the tumor DNA of 20 DLBCL patients after a centralized histopathologic review.Methods: The study included 20 patients having DLBCL. The quality and quantity of tumor cells were accessed by H&E staining and correlated with histopathology and Immunohistochemistry (IHC) status. Patients were grouped as ABC (activated B-cell), PMBL (primary mediastinal large B-cell lymphoma), and other or unclassified subtypes. The lymphoma panel of 133 was designed on targeted sequencing of multiple genes for the coding regions through NGS. The libraries were prepared and sequenced using the Illumina platform. The alignment of obtained sequences was performed using Burrows-Wheeler Aligner and identification of somatic mutations was done using LoFreq (version 2) variant caller. The mutations were annotated using an annotation pipeline (VariMAT). Previously published literature and databases were used for the annotation of clinically relevant mutations. The common variants were filtered for reporting based on the presence in various population databases (1000G, ExAC, EVS, 1000Japanese, dbSNP, UK10K, MedVarDb). A custom read-depth-based algorithm was used to determine CNV (Copy Number Variants) from targeted sequencing experiments. Rare CNVs were detected using a comparison of the test data read-depths with the matched reference dataset. Reportable mutations were prioritized and prepared based on AMP-ASCO-CAP (Association for Molecular Pathology-American Society of Clinical Oncology-College of American Pathologists), WHO guidelines, and also based on annotation metrics from OncoMD (a knowledge base of genomic alterations).Results: The informativity of the panel was 95 percent. NOTCH 1 was the most frequently mutated gene in 16.1% of patients followed by 12.9% who had ARID1A mutations. MYD88 and TP53 mutations were detected in 9.6% of the patient while 6.4% of patients had CSF3R mutations. NOTCH 1 and TP 53 are the most frequently reported gene in the middle age group (40-60). Mutation in MYD88 is reported in every age group. MYD88 (51%) is the most common mutation in ABC subtypes of DLBCL, followed by NOTCH 1 (44%) and SOCS 1 (33%) according to our findings. NOTCH 1 mutations are frequent in ABC and PMBL subtypes. Closer investigation reveals missense mutation is the most frequent mutation observed in the total cohort targeting 68.4% followed by frameshift deletion reported in 26.3%. Six novel variants have been discovered in this study.Conclusions: This study demonstrates the high yield of information in DLBCL using the NGS Lymphoma panel. Results also highlight the molecular heterogeneity of DLBCL subtypes which indicates the need for further studies to make the results of the NGS more clinically relevant.

Hemophagocytic lymphohistiocytosis as an onset of diffuse large B-cell lymphoma: A case report.

A 53-year-old male presented with a 1-month history of hyperpyrexia. The clinical manifestations revealed hemophagocytic lymphohistiocytosis (HLH). Although a lymph node biopsy could not be obtained, a bone marrow biopsy revealed the activated B-cell subtype of diffuse large B-cell lymphoma (DLBCL). After being treated with HLH-1994 (dexamethasone and etoposide), a rituximab-containing chemotherapy and target agents involving bortezomib, the patient achieved remission. To understand the molecular profile of patient, next-generation sequencing and MYD88 L265P mutation examinations were performed, and the patient was determined to be positive for the MYD88 L265P mutation. Reports of DLBCL with plasmacytic differentiation and a MYD88 innate immune signal transduction adaptor L265P mutation concurrent with HLH are rare. Early recognition, precise diagnosis and timely therapy are pivotal in improving patient prognosis. Furthermore, molecular profiling enables researchers to develop potential therapies aimed at the activated NF-κB and endoplasmic reticulum stress signaling pathways. The present study highlights this pathogenesis and provides suggestions for further individualized therapeutics.

Immunohistochemical subtyping of diffuse large B-cell lymphoma into germinal center B-cell and activated B-cell subtype, along with correlation of the subtypes with extranodal involvement, serum lactate dehydrogenase, and positron emission tomography scan-based response assessment to chemotherapy

Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphoma in Indian population and is divided into the prognostically important subtypes, germinal center B-cell (GCB) and activated B-cell-like (ABC), using immunohistochemistry-based algorithm. The present study aims to evaluate the influence of immunohistochemical derived DLBCL subtype, GCB or ABC on prognostically significant variables - extranodal involvement and serum lactate dehydrogenase (LDH) level at presentation, and response to chemotherapy assessed on pre- and posttreatment fluorodeoxyglucose-positron emission tomography study. This was a retro-prospective, 2-year observational study at a tertiary health-care center, New Delhi. The study population includes a total 236 cases of DLBCL. According to the Hans algorithm, DLBCL cases were allocated to the GCB and ABC subgroups. For comparison of mean values, independent t-test and analysis of variance were used. For this purpose, we used SPSS 20.0 software. P < 0.05 was considered as statistically significant. Ninety-eight patients (41.5%) had GCB immunophenotype and 138 patients (58.5%) were ABC. A significant difference was observed between mean baseline level of LDH between GCB and ABC subtypes (P < 0.05). The proportion of cases with extranodal involvement was comparatively higher in ABC subtype (P < 0.05). Association between response to chemotherapy with DLBCL immunophenotypes was found to be highly significant (P < 0.00). The response rates were much better in GCB subtype. The mean baseline level of LDH is significantly higher in ABC subtype. The proportion of cases with extranodal involvement was comparatively higher in ABC and shows poor response to chemotherapy as compared to GCB. Baseline LDH level was found to be important prognostic marker in the DLBCL.

Abstract 1755: IKS03, a CD19-targeted antibody drug conjugate with enhanced efficacy and tolerability for treatment of B-cell lymphomas

Abstract Background: Lymphoma is the most common hematological malignancy with non-Hodgkin B-cell lymphomas (NHL), including those with high unmet clinical need such as relapsed/refractory Diffuse Large B Cell Lymphoma (DLBCL), and Mantle-Cell Lymphoma (MCL), representing 90% of all lymphoma cases. CD19 is an attractive target for antibody-directed lymphoma therapies as it is expressed in most B cell malignancies, with normal tissue expression limited to B-cells thus reducing any on-target toxicity concerns. Antibody Drug Conjugates (ADCs) are the focus of intense interest as a means to provide selective tumor killing with increased efficacy and less toxicity than standard of care chemotherapies. IKS03 is an ADC comprised of an anti-CD19 antibody conjugated to a proprietary DNA-crosslinking PBD prodrug and includes a tumor-selective glucuronide-trigger technology for payload release and activation. ADC activity requires processing by beta-glucuronidase, a lysosomal enzyme often upregulated in tumor cells, while normal tissues with low levels of the enzyme are less able to process the ADC and are differentially spared. Methods: IKS03 was generated via chemo-enzymatic bioconjugation at defined sites on the antibody yielding an ADC with a drug to antibody ratio of 2. In vivo efficacy was evaluated in CD19-expressing cell line-derived xenograft models in mice including Farage (DLBCL, Germinal Centre B-cell subtype), OCI-LY10 (DLBCL, Activated B-Cell subtype), Granta-519 (MCL) and Ramos (Burkitt’s lymphoma). Activity was compared to benchmark ADCs known to have demonstrated clinical efficacy. Efficacy was also assessed in low passage DLBCL patient-derived xenograft models of known genomic profile. Toxicology studies were conducted in cynomolgus monkeys with immunophenotyping by flow cytometry included to quantify the level of normal B-cells following IKS03 administration. Results: IKS03 is highly effective in causing tumor regressions in DLBCL models at doses that are well tolerated. Complete regressions were observed with a single dose of 0.1 mg/kg in the Farage xenograft model. IKS03 is also highly active in the Granta-519 MCL model, with complete regressions observed with a single dose of 0.1 mg/kg. Tumor regression was observed in a high-grade triple-hit lymphoma PDX model with a single 0.3 mg/kg dose. IKS03 was tolerated in monkeys at the highest dose tested of 1.5 mg/kg (single dose). IKS03 cross reacts with monkey CD19 and reduced B-cell depletion was observed in relation to comparator agents, even at doses much greater (1.5 mg/kg) than that needed for complete responses in B-cell lymphoma mouse models (0.1 mg/kg). Conclusion: Preclinical data demonstrates that IKS03’s advanced ADC design results in an increased therapeutic margin compared to traditional ADCs with DNA-crosslinking payloads, with increased efficacy and decreased toxicity. Citation Format: Jenny Thirlway, Adam Lodge, Daniel J. Williamson, Justyna Mysliwy, Jutta Deckert, Xavier Chauchet, Laura Cons, Yun-Hee Park, Hyun-Min Ryu, Na Ra Han, Ho Young Song, Chul-Woong Chung, Robert J. Lutz. IKS03, a CD19-targeted antibody drug conjugate with enhanced efficacy and tolerability for treatment of B-cell lymphomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1755.

Anticancer activity of IRAK-4 inhibitors against canine lymphoid malignancies.

The interleukin-1 receptor-related kinase 4 (IRAK4), downstream of myd88, plays an essential role in hyperactive TLR signalling seen in some B-cell lymphomas. In particular, efficient IRAK4 inhibitors of activated B-cell subtype of human diffuse large B-Cell lymphoma (DLBCL) are being developed. However, the anticancer effect of IRAK-4 inhibitors in veterinary medicine has not been elucidated. It is therefore explored in this study involving the GL-1 and CL-1 canine lymphoma cell lines in vitro. MyD88 expression was analysed using polymerase chain reaction. GL-1 and CL-1 cells were subjected to concentration- and time-dependent treatment with an IRAK-4 inhibitor and assessed for viability, TLR signalling association and apoptosis using a cell counting Kit-8 assay, Western blotting and flow cytometry. The GL-1 and CL-1 cells exhibited enhanced MyD88 expression, however, canine peripheral blood mononuclear cells (cPBMCs) did not. The IRAK-4 inhibitor reduced cell viability in a dose- and time-dependent manner, significantly reduced the phosphorylation of molecules associated with TLR signalling at IC50 such as IRAK1, IRAK4, NF-κB and STAT3, and induced apoptosis in GL-1 and CL-1 cells. The anticancer effect of the IRAK-4 inhibitor on canine lymphoma cells is mediated by apoptosis via downregulation of TLR signalling.

Cost-Effectiveness of Molecularly Guided Treatment in Diffuse Large B-Cell Lymphoma (DLBCL) in Patients under 60.

Simple SummaryDiffuse large B cell Lymphoma (DLBCL) can be categorized into cell of origin (COO) subtypes. Genetic tests are used to detect which subtype of DLBCL a patient has. One subtype of DLBCL, activated B-cell like (ABC), is associated with a comparatively poorer prognosis. New evidence suggests that patients under 60 with ABC-DLBCL may benefit from ibrutinib added to their standard care treatment regimen. Genetic testing and treatments like ibrutinib can be costly to publicly funded healthcare systems. Here, we report a cost-effectiveness analysis of the addition of Ibrutinib to treat patients under the age of 60 with ABC-DLBCL. Our analysis found that the use of genetic testing to diagnose ABC-DLBCL and providing ibrutinib along with standard care treatment has a low to moderate probability of being cost-effective, depending on decision maker willingness to pay and the cost of the genetic test.Background: Classifying diffuse large B-cell lymphoma (DLBCL) into cell-of-origin (COO) subtypes could allow for personalized cancer control. Evidence suggests that subtype-guided treatment may be beneficial in the activated B-cell (ABC) subtype of DLBCL, among patients under the age of 60. Methods: We estimated the cost-effectiveness of age- and subtype-specific treatment guided by gene expression profiling (GEP). A probabilistic Markov model examined costs and quality-adjusted life-years gained (QALY) accrued to patients under GEP-classified COO treatment over a 10-year time horizon. The model was calibrated to evaluate the adoption of ibrutinib as a first line treatment among patients under 60 years with ABC subtype DLBCL. The primary data source for efficacy was derived from published estimates of the PHOENIX trial. These inputs were supplemented with patient-level, real-world data from BC Cancer, which provides comprehensive cancer services to the population of British Columbia. Results: We found the cost-effectiveness of GEP-guided treatment vs. standard care was $77,806 per QALY (24.3% probability of cost-effectiveness at a willingness-to-pay (WTP) of $50,000/QALY; 53.7% probability at a WTP of $100,000/QALY) for first-line treatment. Cost-effectiveness was dependent on assumptions around decision-makers’ WTP and the cost of the assay. Conclusions: We encourage further clinical trials to reduce uncertainty around the implementation of GEP-classified COO personalized treatment in this patient population.

Frequency of Diffuse Large B- Cell Lymphoma, Activated B- Cell Type

Background: Diffuse large B-cell lymphoma (DLBCL) is a heterogenous neoplasm of lymphoid series cells constituting 30%–40% of non-Hodgkin lymphoma in adults. DLBCL presents significant heterogeneity Aim: To determine the frequency of activated B-cell subtype of DLBCL by using Hans algorithm. Methods: It was designed as a cross sectional study conducted in a duration of six months (5th Jun 2018 to 7th January 2019). All data was collected by using a Performa. The clinical parameters like age and gender were recorded. The histopathological method used was paraffin embedding and hematoxylin–eosin staining. The immunohistochemical technique used on serial sections was immune-enzymatic soluble complex method. SPSS version 23 was used for data entry and analysis. Mean and standard deviation was determined for quantitative variables like age and tumor size. Calculation of qualitative variables like gender, tumor site (nodal as well as extra-nodal) and immunohistochemical staining was done in the form of frequencies and percentages. The mean age of the patients was 48.55±15.41 years. Majority of them were male with frequency of 69(71.9%) and female were 27(28.1%). Mean size of lymphoma was 12.44±4.24cm in the patients. Most of the cells were nodal with frequency of 58(60.4%) and a few extra nodal with frequency of 38(39.6%). It was noticed that germinal centre B-cell type was in 53(55.2%) and activated B-cell type lymphoma was present in 43(44.8%). Conclusion: It is observed that frequency of the activated B–cell type lymphoma are less common in patients with lymphoma. Keyword: Lymphoma, Germinal, carcinoma

Primary Adrenal Lymphoma: When B Symptoms Appear Hand in Hand with Adrenal Masses.

Primary adrenal lymphoma (PAL) is a very rare type of non-Hodgkin’s lymphoma (NHL). Herein, we report a case of NHL of both adrenal glands in a 69-year-old man. The patient was admitted because of a 1-month history of B symptoms and symptomatic hypotension. Biochemical analysis showed normocytic normochromic anaemia, hyponatraemia, hyperkalaemia, and elevated lactate dehydrogenase, C-reactive protein and D-dimers. A computed tomography scan revealed bilateral enlargement of the adrenal glands. There was no evidence of endocrine adrenal dysfunction. The mass in the right adrenal gland was biopsied and histopathology identified a diffuse large B-cell lymphoma of the activated B-cell subtype. A positron emission tomography 18F-fluorodeoxyglucose scan showed intensive hypermetabolic lesions involving both adrenal glands, as well as other locations, with higher uptake in the adrenal glands. Taken together, these findings suggested the diagnosis of PAL. The patient responded favourably to debulking therapy and is currently undergoing chemotherapy.LEARNING POINTSPrimary adrenal lymphoma is a rare condition presenting with unspecific symptoms; diagnosis requires histopathological confirmation.Adrenal function must be evaluated to rule out insufficiency.Positron emission tomography may reveal hitherto unsuspected extension of disease and should be performed where available.