Diffuse Large B-cell Research Articles

Novel Therapies for Relapsed or Refractory Diffuse Large B-Cell Lymphoma.

The most common type of non-Hodgkin lymphoma in adults is diffuse large B-cell (DLBCL). There is a historical unmet need for more effective therapies in the 2nd and 3rd line setting. Emerging immunochemotherapies have shown activity in small studies of heavily pre-treated patients with prolonged remissions achieved in some patients. Anti-CD19 CAR (chimeric antigen receptor) T cells are potentially curative in the 3rd line and beyond setting and are under investigation in earlier lines of therapy. Antibody-drug conjugates (ADC’s) such as polatuzumab vedotin targeting the pan-B-cell marker CD79b has proven effectiveness in multiply-relapsed DLBCL patients. Tafasitamab (MOR208) is an anti-CD19 monoclonal antibody producing prolonged remissions when combined with Lenalidomide (LEN) in patients who were not candidates for salvage chemotherapy or autologous stem cell transplant. Selinexor, an oral, small-molecule selective inhibitor of XPO1-mediated nuclear export (SINE), demonstrated prolonged activity against heavily-pretreated DLBCL without cumulative toxicity and is being investigated as part of an oral, chemotherapy-free regimen for relapsed aggressive lymphoma. This article reviews current strategies and novel therapies for relapsed/refractory DLBCL.

Pharmacologic Targeting MCL1 with AZD5991 Induces Apoptosis and Mitochondrial Dysfunction in Non-Hodgkin Lymphoma (NHL) Cells

BCL2 family proteins determine cell fate and comprise pro-apoptotic "initiators" (NOXA, BIM, PUMA), anti-apoptotic "guardians" (BCL2, MCL1, BCLX) and pro-apoptotic effectors (BAX/BAK). Venetoclax, a BCL2 inhibitor, received regulatory approval in therapy of chronic lymphocytic leukemia (CLL) and acute myeloid leukemia. However, venetoclax is modestly effective in NHL. MCL1 is a short-lived pro-survival protein that is frequently overexpressed in NHL, leading to increased cell survival and drug resistance. MCL1 transgenic mice develop lymphoma, mostly of the diffuse large B-cell (DLBCL) type. Thus, selective targeting MCL1 represents a promising pharmacologic strategy in NHL. AZD5991 is a small molecule inhibitor highly selective towards MCL1 (Tron et al, 2018) in clinical trials. Here we explored pre-clinical activity of AZ5591 in NHL. Experiments were conducted in activated B-cell (ABC)-like (OCI-LY3, U-2932) and germinal center (GC)-like (OCI-LY18/19, SUDHL4/6/10, VAL) DLBCL cell lines; parental and ibrutinib-resistant Mino mantle cell lymphoma (MCL) cell lines. Peripheral blood mononuclear cells were isolated from patients with CLL and MCL and co-cultured with BAFF- or CD40L-expressing stroma. AZD5991 was obtained from Activ Biochem Ltd. MCL1 expression was detected in 10 tested DLBCL cell lines, highly expressed in 7/10. Meanwhile, BCLX was upregulated in eight, and BCL2 in 5/10 lines. Four cell lines expressed all three proteins. To confirm relevance of this data, we conducted analysis of primary DLBCL lymph nodes (n=30). MCL1 and BCLX were expressed in 50% and 80% of GC-like, and 10% and 25% of non-GC tumors, respectively. Treatment with AZD5991 restricted growth of DLBCL cells in a dose-dependent manner. GC-like cell lines VAL, SU-DHL4 and SU-DHL6 were highly susceptible with IC50~0.2 µM. Interestingly, they expressed relatively low levels of MCL1. Similarly, parental and ibrutinib-resistant Mino MCL cells were susceptible to MCL1 inhibition, with IC50 of 0.1 µM and 0.5 µM, respectively. Meanwhile, SU-DHL10, OCI-LY19 (GC-like), OCI-LY3 and U-2932 (ABC-like) cells were resistant to AZD5591. Consistent with its mechanism of action, immunoprecipitation assays showed that AZD5591 displaced BIM from MCL1 in NHL cells. We then evaluated MCL1 inhibition in primary neoplastic B-cells. CLL and MCL cells from patients were co-cultured with either CD40L- or BAFF-expressing stroma for 24 h. While CD40L predominantly induced BCLX, BAFF upregulated MCL1 in those cells. Consistent with this, BAFF-stimulated cells were highly sensitive to AZD5991, while CD40L-stimulated cells exhibited resistance. Since MCL1 acts in balance with pro-apoptotic effectors BAX and BAK at the mitochondrial membrane, we assessed the physiological effects of MCL1 inhibition on the mitochondrial function. Treatment with AZD5991 induced mitochondrial depolarization and led to a reduction in mitochondrial mass as well as increased generation of reactive oxygen species. This was accompanied by a decrease in cellular maximal respiratory capacity in both DLBCL and parental/ibrutinib-resistant MCL cells. Meanwhile, the rate of glycolysis was not significantly impacted. Interestingly, MCL1 inhibition induced mitophagy in sensitive (VAL) but not resistant cells (OCI-LY3). Next, we evaluated AZD5991 for synthetic lethality in a functional MTS-based screening assay using a panel of 189 small molecule inhibitors that target a variety of distinct signaling pathways activated in cancer (Tyner et al, 2018). AZD5991 demonstrated synergy with other BH3-mimetics. Co-treatment of DLBCL cells with BCL2/X inhibitors AZ4320, venetoclax and navitoclax overcame resistance to AZD5991. In summary, MCL1 inhibition using selective BH3-mimetic AZD5991 restricts cell proliferation and induces apoptosis in a subset of DLBCL, ibrutinib-resistant MCL cells and primary neoplastic B cells. MCL1 inhibition leads to mitochondrial dysfunction and mitophagy. Resistance to MCL1 inhibition may be overcome by concurrent targeting of alternative anti-apoptotic proteins (BCL2/X) in NHL. Disclosures Tyner: Syros: Research Funding; Gilead: Research Funding; Takeda: Research Funding; Aptose: Research Funding; Constellation: Research Funding; AstraZeneca: Research Funding; Array: Research Funding; Janssen: Research Funding; Incyte: Research Funding; Genentech: Research Funding; Seattle Genetics: Research Funding; Petra: Research Funding; Agios: Research Funding. Danilov:Abbvie: Consultancy; Celgene: Consultancy; Rigel Pharmaceuticals: Consultancy; Bristol-Myers Squibb: Research Funding; Aptose Biosciences: Research Funding; Astra Zeneca: Consultancy, Research Funding; Verastem Oncology: Consultancy, Research Funding; Takeda Oncology: Research Funding; Gilead Sciences: Research Funding; Bayer Oncology: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; TG Therapeutics: Consultancy; Nurix: Consultancy; BeiGene: Consultancy; Pharmacyclics: Consultancy; Karyopharm: Consultancy.

Oral Nanatinostat (Nstat) and Valganciclovir (VGCV) in Patients with Recurrent Epstein-Barr Virus (EBV)-Positive Lymphomas: Initial Phase 2 Results

Oral Nanatinostat (Nstat) and Valganciclovir (VGCV) in Patients with Recurrent Epstein-Barr Virus (EBV)-Positive Lymphomas: Initial Phase 2 Results

Longitudinal Serological Response to Sars-COV-2 in Patients Affected By Hematologic Diseases

Longitudinal Serological Response to Sars-COV-2 in Patients Affected By Hematologic Diseases

Borrelia burgdorferi in primary cutaneous lymphomas: a systematic review and meta-analysis.

The association between Borrelia burgdorferi and primary cutaneous lymphoma is still unclear. This systematic review and meta-analysis aims to define the association of Borrelia burgdorferi with primary cutaneous lymphoma and its different entities. Electronic databases were searched for all studies that assessed the presence of Borrelia burgdorferi DNA in specimens of primary cutaneous lymphoma. The association between Borrelia and primary cutaneous lymphomas was assessed with an odds ratio (significant p<0.05); cutaneous specimens with no lymphoproliferative disorders were used as controls. A secondary analysis was performed to assess the prevalence of Borrelia infection in different lymphoma entities. Ten studies with 506 primary cutaneous lymphomas and 201 controls were included. The prevalence of Borrelia DNA positivity was highly heterogeneous among studies from different regions. Borrelia DNA positivity was significantly associated with primary cutaneous lymphomas (odds ratio = 10.88; p<0.00001). The prevalence of Borrelia DNA positivity was similar among different entities (marginal zone: 7.3%; follicular: 8.1%; diffuse large B-cell: 7.5%; mycosis fungoides: 8%). Borrelia burgdorferi is significantly associated with primary cutaneous lymphomas, with no differences among the several lymphoma entities (both B-cell and T-cell), but with strong geographical differences. Molecular testing for Borrelia would be justified in patients with primary cutaneous lymphoma from endemic areas.

S2941 Ulcerated Gastric Large Diffuse B-Cell Lymphoma in a 95-Year-Old Female

INTRODUCTION: Gastric lesions are commonly benign and include mucosal or mesenchymal tumors. Malignant lesions include adenocarcinoma, lymphoma, stromal and neuroendocrine tumors. Primary gastric lymphomas are an uncommon etiology for gastric lesions. We describe a case of primary gastric Diffuse large B-Cell lymphoma in a patient with concomitant myelodysplastic syndrome. CASE DESCRIPTION/METHODS: The patient is a 95 year old female with a history of myelodysplastic syndrome that presented to the hospital for syncope. Labs on presentation showed a hemoglobin of 5.4, WBC 15.4, platelet 837, BUN 56, and Creatinine 1.36. The patient was transfused two units of packed red blood cells and her hemoglobin increased to 8.3. Rectal exam was performed revealing heme-positive stool with reported episodes of melena at home. Twice daily pantoprazole was initiated. She underwent endoscopic evaluation demonstrating an ulcerated gastric lesion within the stomach body. Biopsy was performed resulting in atypical/neoplastic lymphocytes infiltrating the gastric lamina propria consistent with diffuse large B cell lymphoma. Immunostains were positive for CD45, CD20, PAX5, MUM1, BCL2, c-myc, BCL6 and CD30. The patient’s hemoglobin remained stable and she was discharged to follow up with oncology. One month after discharge, the patient denied wanting to pursue systemic therapy and underwent external beam radiation. The patient required multiple blood transfusions in the following months and ultimately pursued chemotherapy with R-CHOP five months following her diagnosis. DISCUSSION: Primary gastric lymphomas are rare, representing 0.9% of gastrointestinal tumors and up to 5% of gastric malignancies. It is considered an extranodal form of Non-Hodgkin's lymphoma. Large diffuse B-cell lymphoma is one common subtype and according to the National Cancer Institute, at time of diagnosis, the majority of patients have stage 4 disease severity with a corresponding 5 year survival rate of 52.7%. Presenting symptoms may be nonspecific and can mimic other disorders such as peptic ulcer disease or gastritis. Up to 30% of patients report gastrointestinal bleeding as a presenting or associated symptom. Mainstay treatment is 3–4 rounds of R-CHOP therapy with or without radiation. Surgery is reserved for patients with perforation, hemorrhage or obstruction. Our patient presented with symptomatic anemia inducing a syncopal episode likely form gastrointestinal blood loss. As of now, the patient has had a total of 2 cycles of R-CHOP therapy.Figure 1.: Ulcerated mass in the stomach body.Figure 2

The cytopathological spectrum of lymphomas in effusions in a tertiary center in Taiwan.

We retrospectively reviewed effusion lymphomas from our archives in a tertiary center from July 2011 to June 2019. We identified 59 specimens from 43 patients, including 7 cases with primary effusion lymphoma (PEL) and 36, secondary effusion involvement. Half of the secondary cases presented concurrently with effusion lymphoma, while the remaining half-experienced effusion lymphoma during disease progression. All patients with PELs were males with a median age of 77 and presented with massive pleural effusion. None was HIV-related. Two (29%) PEL cases were positive for human herpes virus 8 (HHV8). The only case with plasmablastic phenotype in the PEL group was positive for both HHV8 and EBV. Four patients died shortly after diagnosis; while the remaining three were alive at the last follow-up (two at 13 months and one at 99 months). Of the secondary cases, diffuse large B-cell lymphoma/high grade B-cell lymphoma was the most common (n = 16, 44%), followed by mantle cell lymphoma (n = 5, 14%). Only 8 cases (22%) were T-cell neoplasms. Prognosis for patients with secondary effusion involvement was dismal, with 1- and 2-year overall survival rates at 17% and 8%, respectively. We found a wide cytopathological spectrum of effusion lymphoma in Taiwan. Most of our PEL cases were distinct from that defined in the World Health Organization scheme by a B-cell phenotype, HHV8-negativity, and absence of immunodeficiency. As compared to PEL cases, the prognosis of those with secondary involvement was extremely poor.

Hepatitis C Virus Infection and Treatment as Independent Prognostic Factors in Diffuse Large B-Cell Lymphoma Egyptian Patients.

Egypt is one of the highest hepatitis C virus (HCV) endemic areas. Chronic HCV infection has extra-hepatic manifestations, including non-Hodgkin lymphoma (NHL). Diffuse large B-cell lymphoma (DLBCL) is commonly associated with HCV infection. The prognostic value of HCV infection and HCV treatment in patients with DLBCL remains unclear until now. The aim of our study is to evaluate the impact of HCV infection and HCV treatment as independent prognostic factors on the event-free survival (EFS) and overall survival (OS) in Egyptian patients with HCV associated DLBCL. This study included 353 patients with DLBCL, collected retrospectively. While 34 patients with HCV who received HCV antiviral therapy were collected prospectively. Patient's characteristics were collected from the patient records at the time of diagnosis. The status of the patients about HCV infection and HCV treatment were also recorded. Disease progression, relapse, retreatment or deaths were also verified through medical records. EFS and OS were calculated. EFS and OS significantly decrease in HCV infected and HCV non-treated patients when compared with HCV non-infected and HCV treated patients, respectively. HCV infection but not HCV treatment was independently associated with EFS and OS using univariate and multivariate analysis. Hepatitis C virus infection is an independent prognostic factor for EFS and OS in diffuse large B-cell lymphoma. HCV treatment is associated with higher EFS and OS but can not be considered as an independent prognostic factor.

Abstract 1054: Vaccination history and risk of lymphoma and its major subtypes

Abstract Background Factors associated with chronic immune stimulation such as transplantation, autoimmune disorders and chronic infections have been linked to the development of lymphoma, while atopic conditions have been associated with a lower risk of lymphoma. Vaccinations, in part due to their ability to induce immune responses, have been hypothesized to play a role in lymphoma etiology, but there are few studies with mixed results and limited data on lymphoma subtypes. Methods We used a clinic-based study of 2461 newly diagnosed lymphoma cases and 2253 frequency-matched controls enrolled from 2002-2015. Participants self-reported lymphoma risk factors and history of vaccinations for hepatitis A, hepatitis B, chicken pox, yellow fever and influenza. Pathology was centrally reviewed, and included 644 chronic lymphocytic leukemia, 528 follicular (FL), 414 diffuse large B-cell (DLBCL), 170 marginal zone, 127 mantle cell, 119 T-cell, 181 Hodgkin, and 278 other lymphoma subtypes. We used polytomous logistic regression to estimate odds ratio (OR) and 95% confidence intervals (CI) of vaccinations with lymphoma risk overall and for subtypes, adjusting for age, sex, socioeconomic status, body mass, alcohol use, smoking history, recreational sun exposure, and family history of hematological malignancies. Results The median age at diagnosis for lymphoma cases was 64 years (range: 18-93) and 58% were male; the median age at enrollment for controls was 62 years (range: 18-91) and 53% were male. The most common vaccinations among controls were influenza (90%) followed by hepatitis B (41%), hepatitis A (30%), chicken pox (20%), and yellow fever (11%). Vaccination for influenza was inversely associated with lymphoma risk overall (OR=0.75, CI:0.61-0.93, P=0.01), with an inverse trend for increasing number of vaccinations (P=0.0001). In subtype analyses, Influenza vaccination was inversely associated with risk of FL (OR=0.59, CI:0.43-0.80, P=0.001), DLBCL (OR=0.64, CI:0.44-0.92, P=0.02) and other lymphoma subtypes (OR=0.66, CI:0.43-1.00, P=0.05). Vaccination for hepatitis A was inversely associated with lymphoma risk (OR=0.79, CI:0.65-0.96, P=0.02), with no significant subtype-specific associations. No evidence of association with vaccination for hepatitis B (OR=0.95, CI:0.79-1.14) and chicken pox (OR=0.83, CI:0.68-1.02) for lymphoma risk overall or for any subtypes. Although no evidence of an association with vaccination for yellow fever (OR=0.81, CI:0.62-1.06) for lymphoma risk, it was inversely associated with DLBCL risk (OR=0.42, CI:0.22-0.80). Conclusion Vaccination for influenza and hepatitis A were inversely associated with overall lymphoma risk. DLBCL was associated with vaccination for influenza and yellow fever. FL was associated with vaccination for influenza. Understanding the underlying mechanisms for a putative protective effect of certain vaccines could provide new clues to the etiology and prevention of lymphoma. Citation Format: Geffen Kleinstern, Dennis P. Robinson, Carrie A. Thompson, Melissa C. Larson, Cristine Allmer, Grzegorz S. Nowakowski, Timothy G. Call, Stephen M. Ansell, Andrew L. Feldman, Neil E. Kay, Thomas M. Habermann, Susan L. Slager, James R. Cerhan. Vaccination history and risk of lymphoma and its major subtypes [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1054.

Ophthalmopathologic characterization of multicentric or metastatic neoplasms with an extraocular origin in dogs and cats

To characterize the frequency and distribution of secondary ocular neoplasms in dogs and cats identified during necropsy. A retrospective analysis of necropsy records of dogs and cats was conducted, and cases with metastatic/multicentric neoplasms with an extraocular origin, involving the eyes, were selected. From January 2015 to January 2019, we identified a total of 233 dogs and 100 cats with metastatic disease. Of these, 11.6% (27/233) of the dogs and 13% (13/100) of the cats had ocular metastases. Lymphoma was the most common multicentric neoplasm involving the eyes of both species. In dogs, these neoplasms occurred bilaterally, predominantly in the anterior uvea, and were diffuse large B-cell, T-lymphoblastic, peripheral T-cell not otherwise specified, and lymphocytic B-cell lymphomas. In cats, feline leukemia virus (FeLV)-associated T-cell lymphoma was the most common. Mammary carcinoma was the second most common ocular metastatic neoplasm in bitches, with a predominantly unilateral involvement of the uveal tract. In cats, following lymphoma, pulmonary and squamous cell carcinomas were the most common multicentric/metastatic neoplasms of the eyes. Individual cases of cholangiocarcinoma, hemangiosarcoma, and chemodectoma in dogs, as well as mammary gland cribriform carcinoma, salivary gland carcinoma, and histiocytic sarcoma in cats were detected. The eyes of dogs were mostly affected by lymphoma or mammary gland carcinoma, while those of cats mostly had lymphoma, pulmonary carcinoma, or squamous cell carcinoma as the main metastatic/multicentric neoplasm. To the best of our knowledge, this is the first study to identify intraocular metastatic cholangiocarcinoma and chemodectoma in dogs, aside from identifying salivary gland carcinoma in cats.

Rituximab is not associated with increased risk of second primary malignancies in Israeli patients with diffuse large B cell lymphoma treated with RCHOP regimen

It is unknown whether rituximab increases the risk of second primary malignancies (SPMs) in patients with diffuse large cell B-cell lymphoma (DLBCL). We assessed SPMs in DLBCL patients diagnosed between 1996 and 2014 in comparison with the general Israeli population and dependent on rituximab treatment. Jewish patients had no increased risk for SPMs. Arab-DLBCL females had a higher SPMs rate compared to the general Arab-females population [SIR (95%CI) 1.86 (1.08–2.98)]. Incidence and time to SPMs, in both Jewish and Arab patients, were unaffected by rituximab. Risk for breast and thyroid cancers, in Arab and Jewish females respectively, were higher in the pre-rituximab era [SIR(95%CI) 5.25 (1.41–13.43) and SIR(95%CI) 3.85 (1.41–8.38), respectively]. Age ≥60 years was the only predictor for increased risk of SPM (HR = 2.5, p < .01). The increased risk of SPMs in specific subgroups of patients that were treated in the pre-rituximab era may reflect stringent medical surveillance employed in these populations.

Potential of Circulating Tumor DNA for the Management of Patients With Lymphoma.

The characterization of circulating tumor-derived DNA (ctDNA) has recently emerged in the field of oncology as a powerful method to identify tumor-specific genetic aberrations using peripheral blood testing. Several technical precautions are needed at the pre-analytic stage (given the short half-life of free nucleic acids in plasma), and numerous techniques-with different sensitivities-are available to identify these molecular aberrations, ranging from the detection of single point mutations to extended genetic screening panels. Although a "liquid biopsy" cannot be substituted for the pathological examination of tissue specimens for diagnostic purposes, it can sometimes complement pathology results or serve as a proxy approach for particular lymphoma presentations where biopsies are sometimes difficult to perform. Moreover, ctDNA testing can characterize, at diagnosis or during treatment, mutations that may contribute to the choice of an optimal targeted therapy (such as Bruton tyrosine kinase or EZH2 inhibitors) or detect the emergence of resistance to those therapies. High levels of ctDNA before treatment appear to be correlated with advanced disease stages and prognosis in diffuse large B-cell and follicular lymphomas. Real-time follow-up of ctDNA levels during therapy in several lymphoma subtypes (diffuse large B-cell and Hodgkin lymphomas) has been explored: preliminary studies have demonstrated that this monitoring technique can predict clinical outcomes (end of treatment response and risk of progression after treatment completion) and that this approach may complement the information provided by metabolic imaging assessments. Technical standardization and careful prospective evaluation of the role of ctDNA monitoring in clinical studies represent current important challenges to allowing its application in routine practice.

Low-Cost, Transcriptional Diagnostic to Accurately Categorize Lymphomas in Low- and Middle-Income Countries

Background: The lack of access to adequate pathology services is a critical roadblock for both improvements in health and sustainable development across lower- and middle-income countries (LMICs). We hypothesized that a low-cost, parsimonious gene expression assay using paraffin-embedded biopsies from LMICs could distinguish lymphoma subtypes and guide treatment. Methods: We reviewed all biopsies obtained between 2006-2018 for suspicion o lymphoma at INCAN hospital in Guatemala City. Gold-standard diagnoses were established by immunohistochemistry and FISH then binned into 9 categories: nonmalignant, aggressive B-cell, diffuse large B-cell (DLBCL), follicular, Hodgkin, mantle cell, marginal zone, NK/T-cell, or mature T-cell lymphoma. We established a chemical ligation probe-based assay (CLPA) that quantifies expression of 37 genes by capillary electrophoresis for <$10 USD/sample. To assign bins based on gene expression, 13 models were evaluated as candidate base learners and class probabilities from each model were then used as predictors in an extreme gradient boosting super learner. An additional two-class model was developed to classify DLBCL cell-of-origin (COO). Cases with call probabilities <0.6 were classified as indeterminate. Findings: Assay failure occurred in 60 (8·9%)/670 biopsies and was enriched among Hodgkin lymphomas (24·8%). 560 diagnostic samples were divided into 70% (n=397) training and 30% (n=163) validation cohorts. Overall accuracy for the validation cohort was 86% [95% CI; 80-91%]. After excluding 28 (17%) indeterminate calls, accuracy increased to 94% [95% CI; 89-97%]. Accuracy for a cohort of relapsed/refractory biopsies (n=39) was 79% and 88% after excluding indeterminate cases. Accuracy for DLBCL COO classification compared to the Hans IHC algorithm (n=51) was 80% [95% CI; 67-90%]. Interpretation: Machine-learning analysis of gene expression accurately classifies paraffin-embedded lymphoma biopsies from LMICs. Low-cost, open source assays could transform diagnosis, subtyping, and assessment of therapeutic targets for patients with cancer worldwide. Funding Statement: American Society of Hematology, US State Department, ASCO, LLS, Celgene and NIH Declaration of Interests: T.G., S.L.D. and R.T. are employees of DxTerity Diagnostics. D.M.W. is a co-founder of Travera, Ajax and Root Diagnostics. He receives consulting or advisory board fees from Magnetar, Bantam, ASELL, Ossium, Myeloid Therapeutics, Daiichi Sankyo, and Elstar. He receives research funding from Daiichi Sankyo and Verastem. The remaining authors declare no conflicts-of-interest. Ethics Approval Statement: This study was approved by the Institutional Review Boards of Dana-Farber Cancer Institute and Stanford University and the Ethics Committee of La Liga Nacional Contra el Cancer Research.

Flow Cytometry in the Differential Diagnosis of CD10-Positive Nodal Lymphomas.

Differences between follicular lymphoma (FL) and diffuse large B-cell lymphoma/high-grade B-cell lymphoma (DLBCL/HGBL) by flow cytometry are underexplored. We retrospectively assessed flow cytometry results from 191 consecutive lymph node biopsies diagnosed with FL or DLBCL/HGBL. The only parameters that differed between the 2 groups in the derivation cohort were forward scatter and side scatter (P < 10-6; area under the curve [AUC], 0.75-0.8) and %CD23 (P = .004; area under the receiver characteristic operating curve, 0.64). However, since light scatter characteristics did not distinguish between grade 3 FL and DLBCL/HGBL, we set out to develop a model with high sensitivity for the exclusion of the latter. Several models, including FS and %CD23, were tested, and 2 models showed a sensitivity of >0.90, with negative predictive values of ≥0.95, albeit with low specificity (0.45 to 0.57). Two simple models enable the exclusion of DLBCL/HGBL with a high degree of confidence.

P.319 Intravenous ketamine long-term use in two bipolar depression cases: 1-year follow-up

Hepatitis B (HBV) reactivation induced by chemotherapy is a problem currently encountered in the management of malignancies. HBV reactivation occurs particularly in patients who were not checked for HBV status, and therefore have not undergone antiviral prophylaxis. HBV reactivation may ultimately lead to fulminant liver failure (FLF). Liver transplantation (OLT), the only remaining effective treatment option, is generally denied for subjects with a recent history of malignancy.We described retrospectively three cases of FLF caused by HBV reactivation in two men and one woman undergoing rituximab-containing chemotherapy for malignant lymphomas: follicular, diffuse large B-cell and lymphoplasmacytic types. The two men reactivated after eight cycles of rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone and the one woman after 13 cycles of rituximab monotherapy; their hematologic disease was in remission. All three patients were hepatitis B surface antigen (HBsAg)-positive with high HBV DNA levels. Neither man had been screened for HBV before chemotherapy; the woman had been treated with lamivudine (LAM) experiencing an HBV flare-up due to emergence of LAM resistance. All patients fulfilled King's College criteria for urgent OLT upon admission to the transplant center and underwent an urgent OLT. Their hemato-oncologic prognosis was considered to be favorable. All three patients are alive (54, 46, and 37 months post-transplantation), tumor-free and HBsAg negative on a standard HBV prophylaxis regimen: hepatitis B immunoglobulin and LAM + adefovir or tenofovir.Before chemotherapy appropriate prophylaxis for HBV reactivation should always be administered to at-risk patients. However, if reactivation with FLF occurs, OLT should not be generally denied. The prognosis of the hematologic malignancy should be assessed; OLT should be considered for patients in remission with a favorable long-term prognosis, for our data suggest acceptable survival.

Interaction kinetics with transcriptomic and secretory responses of CD19-CAR natural killer-cell therapy in CD20 resistant non-hodgkin lymphoma.

We investigated the cytolytic and mechanistic activity of anti-CD19 chimeric antigen receptor natural killer (CD19.CAR.NK92) therapy in lymphoma cell lines (diffuse large B-cell, follicular, and Burkitt lymphoma), including rituximab- and obinutuzumab-resistant cells, patient-derived cells, and a human xenograft model. Altogether, CD19.CAR.NK92 therapy significantly increased cytolytic activity at E:T ratios (1:1–10:1) via LDH release and prominent induction of apoptosis in all cell lines, including in anti-CD20 resistant lymphoma cells. The kinetics of CD19.CAR.NK92 cell death measured via droplet-based single cell microfluidics analysis showed that most lymphoma cells were killed by single contact, with anti-CD20 resistant cell lines requiring significantly longer contact duration with NK cells. Additionally, systems biology transcriptomic analyses of flow-sorted lymphoma cells co-cultured with CD19.CAR.NK92 revealed conserved activation of IFNγ signaling, execution of apoptosis, ligand binding, and immunoregulatory and chemokine signaling pathways. Furthermore, a 92-plex cytokine panel analysis showed increased secretion of granzymes, increased secretion of FASL, CCL3 and IL10 in anti-CD20 resistant SUDHL-4 cells with induction of genes relevant to mTOR and G2/M checkpoint activation were noted in all anti-CD20 resistant cells co-cultured with CD19.CAR.NK92 cells. Collectively, CD19.CAR.NK92 was associated with potent anti-lymphoma activity across a host of sensitive and resistant lymphoma cells that involved distinct immuno-biologic mechanisms.

Tumor ADAM10/ADAM17-Mediated PD-L1 Loss May Predict Poor Outcomes in Diffuse Large B Cell Lymphoma

Introduction Tumor surface matrix metalloproteases ADAM10 and ADAM17 are associated with poor outcomes in multiple malignancies. We previously showed that these proteases shed PD-L1 from lymphoma cell line Karpas-299 to produce soluble PD-L1 (sPD-L1) that induces CD8+ T cell apoptosis (1). While higher levels of soluble PD-L1 are associated with worse prognosis in patients with non-Hodgkin lymphoma treated with standard chemoimmunotherapy (cutoff of 1.52 ng/ml, 3-year OS of 76% versus 89%) (2), it is unknown whether the loss of PD-L1 on the surface of B cell lymphoma cells affects clinical outcomes. We hypothesized that tumors expressing low PD-L1 levels despite high PD-L1 mRNA levels would (1) produce higher ADAM10 or ADAM17 transcript levels and (2) predict poorer prognosis in diffuse large B cell lymphomas. Methods We queried the cancer genome atlas (TCGA) public database for cases of diffuse large B cell lymphoma (DLBCL) for which PD-L1 (CD274) protein levels are reported by reverse phase protein array (RPPA). We then queried TCGA for RNA-Seq data for CD274, ADAM10, and ADAM17 expressed as transcripts per million (FPKM). We re-normalized RPPA data and calculated a PD-L1 protein-to-mRNA ratio for each tumor sample. We then compared groups of high and low PD-L1 protein-to-mRNA ratios for ADAM10 and ADAM17 mRNA expression. We further compared groups of high and low PD-L1 protein-to-mRNA ratios for survival (adjusted for age at diagnosis). All statistical analyses were performed using R Statistical Software (R Foundation). Unpaired student's t-test assessed statistical differences in experimental groups except where otherwise indicated. Figures comprise box plots showing quartile values and individual data points. P&lt;0.05 was considered statistically significant. In figures, p values are denoted &lt;0.05 with *, &lt;0.01 with **, and &lt;0.001 with ***. Results Data were available for a total of 33 diffuse large B cell (DLBC) lymphomas. 18 of those samples were categorized as having low PD-L1 protein-to-mRNA ratios (cutoff 6.38e-6) with the remaining categorized as high ratio. DLBCL tumors demonstrating low PD-L1 protein-to-mRNA ratios expressed significantly higher ADAM10 (p=0.0237) and ADAM17 (p&lt;0.0001) transcripts than tumors expressing low PD-L1 protein-to-mRNA ratios (Figure 1). Furthermore, patients with tumors demonstrating low PD-L1 protein-to-mRNA ratios experienced significantly poorer survival according to likelihood ratio (6.6, p=0.01) and log rank (4.67, p=0.03) tests (Figure 2). Conclusions Our findings may explain a crucial phenomenon seen in diffuse large B cell lymphoma, namely that many purportedly PD-L1-positive tumors appear to express PD-L1 but do not respond to PD-(L)1 inhibitor therapy (3,4). This may be caused by the activity of ADAM10 and ADAM17 to cleave PD-L1, which we previously showed in lymphoma cell line Karpas-299 (1). This posits pre-treatment ADAM10/ADAM17 inhibition to "boost" PD-(L)1 inhibitor therapy in lymphoma. Our results contrast with previous findings that PD-L1-positive DLBCL progression free survival is inferior to PD-L1-negative DLBCL (5). This may be due to different methods of protein detection (immunohistochemistry versus RPPA) and the lack of localization of PD-L1 expression in tumor cells versus the microenvironment. Notably, this study is only correlative with a limited case number. Larger prospective studies will be needed to further elucidate this relationship. Acknowledgments The results shown here are in whole or part based upon data generated by the TCGA Research Network: https://www.cancer.gov/tcga.

Genomic Analysis of R2CHOP-Treated DLBCL Reveals a High-Risk Population Driven By Inflammatory Pathways

Genomic Analysis of R2CHOP-Treated DLBCL Reveals a High-Risk Population Driven By Inflammatory Pathways

Targeting B-Cell Maturation Antigen (BCMA) with CC-93269, a 2+1 T Cell Engager, Elicits Significant Apoptosis in Diffuse Large B-Cell Lymphoma Preclinical Models

B-cell maturation antigen (BCMA), a member of the TNF receptor superfamily, serves as the cell surface receptor for B-cell activating factor (BAFF). Upon binding of BAFF to BCMA, an intracellular signaling cascade is initiated resulting in upregulation of JNK pathway signaling events and NFkB mediated transcription. The canonical expression pattern of BCMA begins on germinal center B-cells and becomes maximally expressed on mature cells such as plasma cells. Given the high degree of expression of BCMA on multiple myeloma (MM) cells, the malignant counterpart to plasma cells, it has become a target of interest for CAR T and antibody mediated modalities such as antibody drug conjugates and bispecific molecules. Recent clinical data from clinical trials employing BCMA targeted CAR T cells or T cell engager (TCE) antibodies have demonstrated significant responses in heavily pretreated myeloma patients with overall response rates ranging from 70% to 80%. As BCMA is known to be expressed in earlier B-cell lineages, we sought to explore the expression of BCMA in non-Hodgkin lymphoma (NHL) and its sensitivity to CC-93269, a 2+1 TCE currently being clinically investigated in MM. NHL is a heterogeneous collection of lymphomas that can be classified into two major subgroups; aggressive lymphomas of which diffuse large B-cell (DLBCL) is the most prevalent subtype and indolent lymphomas of which follicular lymphoma is the largest constituent. We first sought to directly quantitate cell surface expression of BCMA utilizing a flow cytometry system based on a logarithmic dilution of phycoerythrin beads of a known quantity. In a panel of 43 NHL cell lines, we determined that BCMA expression ranged from 43 to 17,048 molecules per cell (median, 420). An isogenic pair of K562 that is null for BCMA expression and K562 constitutively overexpressing BCMA (K562-BCMA) (15,866 molecules/cell) served as negative and positive controls, respectively. Additionally, the MM cell line H929 was profiled to serve as an additional control with a BCMA expression level of 7,065 molecules/cell. Next, utilizing quantitative PCR we found that relative BCMA mRNA expression in the lymphoma cell lines ranged from 0.001 to 0.17-fold when normalized to the H929 MM cell line. Furthermore, we were able to determine that in the lymphoma cells there is a poor correlation between protein expression (mean fluorescent intensity) and mRNA expression (r2, 0.33). We next examined if there was any correlation between BCMA surface expression and T-cell mediated cytotoxicity after administration of CC-93269 in a co-culture assay. We selected 11 DLBCL cell lines with a surface expression ranging from 45 molecules to 17,000 molecules per cells and incubated them in a co-culture system with a defined 1:5 target:effector ratio with CC-93269 (0-200 ng/ml) for five days. Significant apoptosis as measured by annexin V and ToPro-3 staining of CFSE positive target cells was observed in 10 of the 11 cell lines profiled with an IC50 of 0.1 to 38 ng/ml for CC-93269. As controls, the K562 isogenic pair were also profiled with the K562-BCMA cell line exhibiting an IC50 of 0.5 ng/ml and no activity observed against the parental K562 cell line. Additionally, a bispecific antibody where the two binding domains for BCMA was altered to target HEL (hen egg lysozyme) demonstrated no activity against any of the cell lines profiled at a defined dose of 200 ng/ml. No association between CC-93269 activity and BCMA expression or cell of origin was found. To determine the expression of BCMA in primary DLBCL biopsy samples, we developed a novel monoclonal BCMA immunohistochemistry antibody (clone: G12). The antibody and IHC staining protocol were validated to have good on-target specificity in both cell lines and tissues, including MM and DLBCL biopsies, with a range of stain intensity (1-3+) observed in both the golgi and on the plasma membrane. A proof of concept study on a cohort of 110 commercial DLBCL samples is currently underway. Cumulatively, our data demonstrate that BCMA is expressed on the cell surface of a broad panel of NHL cell lines and in primary DLBCL lymph node biopsies. Additionally, the expression levels of BCMA in these preclinical cell line models was sufficient to elicit significant CC-93269 mediated cytotoxicity. These data highlight the potential for the treatment of DLBCL patients with a 2+1 T-cell engager targeting BCMA. Disclosures Hagner: Celgene Corporation: Employment, Equity Ownership, Patents &amp; Royalties. Waldman:Celgene: Employment, Equity Ownership, Patents &amp; Royalties. Gray:Celgene: Employment, Equity Ownership. Yura:Celgene: Employment, Equity Ownership. Hersey:Celgene: Employment, Equity Ownership. Chan:Celgene: Employment, Equity Ownership. Zhang:Celgene: Employment, Equity Ownership. Boss:Celgene Corporation: Employment, Equity Ownership. Gandhi:Celgene Corporation: Employment, Equity Ownership, Patents &amp; Royalties.

The Addition of Ibritumomab Tiuxetan (IT) to the ATG/TLI Preparative Regimen Is Safe and May Enhance Post-Transplant Response and Outcomes for Patients with Relapsed or Refractory B Cell Non-Hodgkin Lymphomas

The Addition of Ibritumomab Tiuxetan (IT) to the ATG/TLI Preparative Regimen Is Safe and May Enhance Post-Transplant Response and Outcomes for Patients with Relapsed or Refractory B Cell Non-Hodgkin Lymphomas