An Artificial Intelligence (AI) Approach for Automated Flow Cytometric Diagnosis of B-Cell Lymphoma

Abstract Background: Multi-parameter flow cytometry (MFC) data is a diagnostic keystone for hematologic malignancies. Data analysis and interpretation requires resources and expertise, full automation is desired. Combination of automated gating strategies and machine learning on MFC data proved effective in detecting AML cell populations vs normal (Aghaeepour et al., 2013). Aim of our approach was to create tools capable of comprehensively analyzing and classifying MFC data. We developed a proof of principle framework to process MFC data generated from samples with suspected B-cell lymphomas. Methods: MFC data was obtained from routine diagnostics of 16,384 patients as follows: normal controls (n=8,493), chronic lymphocytic leukemia (CLL, n=3,412), CLL with increased prolymphocytes (CLL/PL, n=603), follicular lymphoma (FL, n=219), hairy cell leukemia (HCL, n=193), lymphoplasmacytic lymphoma (LPL, n=629), mantle cell lymphoma (MCL, n=269), marginal zone lymphoma (MZL, n=979), monoclonal B-cell lymphocytosis (MBL, n=1,480). Two 9-color combinations of monoclonal antibodies were applied to analyze surface expression of the following antigens besides scatter signals: FMC7, CD10, IgM, CD79b, CD20, CD23, CD19, CD5, CD45, Kappa, Lambda, CD38, CD25, CD11c, CD103, CD22. For dimensionality reduction we used an unsupervised clustering approach (Kohonen et al., 1990). We first generated a consensus self-organizing map (CS) of 100 nodes including all MFC events of multiple preselected cases (5 samples/cohort). For a test case the fluorescent profile of a cell was assigned to its nearest node in the CS (upsampling). We used the resulting distribution for classification by a sequential neural network (NN). Computationally intensive processing steps were run using Amazon Web Services. Mean accuracies were calculated counting individual cases across the entire dataset. All accuracies are described for 10 CS generations and 10-fold cross validation in the classification process. Approaches of data processing were sequentially developed aiming at maximization of classification accuracy. Results: First, we assessed the performance in a binary classification, i.e. the merged group of CLL and MBL (n=4,877) vs. normal (n=8,493) achieving 97% accuracy. Secondly, we applied our approach to the classification of all 9 classes with an accuracy of 74%. Visual inspection of tSNE (van der Maaten et al., 2008) transformed upsampling information revealed good separability between CLL and normal cases but substantial overlap between other lymphoma subtypes (Figure). The following approaches were applied to improve classification: 1) Automated pregating of lymphocytes (CD45/SSC) by DBSCAN analysis (Ester et al., 1996) improved classification. Applying this to both CS generation and upsampling the accuracy of the 9-class problem increased to 78% (p<0.001). 2) Using a CS trained only on cases with unequivocal phenotypes without pregating was significantly better (p=0.01), but pregating overrode this effect (p=0.5). 3) According to the partial overlap of immunophenotypes hierarchical clustering of misclassifications showed overlap mainly between MCL and CLL/PL as well as between LPL and MZL. We therefore changed our approach merging the groups CLL/MBL, MCL/CLL/PL and LPL/MZL while keeping FL, HCL and normal controls separate. Automated classification of these 6 classes resulted in 86% accuracy. 4) Further regrouping into 3 classes, according to the commonly applied routine approach of distinguishing normal cases from CD5+ (CLL, MBL, MCL, CLL/PL) and CD5- B-cell lymphoma (FL, LPL, MZL, HCL) resulted in a classification accuracy of 89%. 5) The misclassified cases in the 3-class experiment had an average infiltration of 10% versus 41% for correctly classified ones. Limiting the automated classification to cases with ≥5% infiltration increased the classification accuracy to 95%. Conclusion: We created a framework for automated classification and analysis of routine diagnostic MFC data. Definition of entities and preselection of cell populations are essential for optimizing classification accuracy. Our approach is the first application of computational MFC and AI on a large scale dataset and proves the feasibility in a routine diagnostic work flow. To further improve the accuracy of classification a future algorithm will integrate distance metrics following SOM generation into the NN classifier. Disclosures Höllein: MLL Munich Leukemia Laboratory: Employment. Schabath:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Download Full-text

TGR-1202, a Novel Once Daily PI3Kδ Inhibitor, Demonstrates Clinical Activity with a Favorable Safety Profile, Lacking Hepatotoxicity, in Patients with Chronic Lymphocytic Leukemia and B-Cell Lymphoma

Blood ◽

10.1182/blood.v124.21.1984.1984 ◽

2014 ◽

Vol 124 (21) ◽

pp. 1984-1984 ◽

Cited By ~ 14

Author(s):

Howard A. Burris ◽

Manish R. Patel ◽

Danielle M. Brander ◽

Owen A. O'Connor ◽

Changchun Deng ◽

...

Keyword(s):

B Cell ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Hematologic Malignancies ◽

Tumor Burden ◽

Lymphocytic Leukemia ◽

Clinical Activity ◽

Employment Equity ◽

Previous Formulation ◽

Equity Ownership

Abstract Background: TGR-1202 is a novel oral, next generation PI3Kδ inhibitor which notably lacks the hepatotoxicity associated with other PI3Kδ inhibitors. Preliminary data from an ongoing Ph I study of TGR-1202 demonstrated clinical activity in patients with advanced hematologic malignancies (ASCO 2014). Herein we present updated results from this Phase I, first in human study of TGR-1202 in patients with relapsed and/or refractory CLL and B-cell lymphoma. Methods: TGR-1202 is administered orally once daily following a 3+3 dose escalation design. Previously treated patients with an ECOG PS ≤ 2 and confirmed diagnosis of B-cell non-Hodgkin lymphoma (NHL), chronic lymphocytic leukemia (CLL), or other lymphoproliferative disorders are eligible. Endpoints include safety, PK/PD, and efficacy. Results: 49 patients have been enrolled to date of various lymphoma subtypes including CLL, follicular lymphoma (FL), Hodgkin’s lymphoma (HL), DLBCL, mantle cell lymphoma (MCL), and marginal zone lymphoma (MZL). Demographics: 76% male, ECOG 0/1/2: 17/31/1, median age of 59 yrs (range: 22-85), median prior treatment regimens: 3 (range: 1-14), and 43% were refractory to prior treatment. 35 patients have been treated at doses ≥ 800 mg of a previous formulation where a threshold effect in activity was observed, and 6 have been treated with an improved micronized formulation (≥ 200 mg). TGR-1202 was well tolerated and no MTD has been reached to date. The only Gr≥3 AE occurring in >5% of patients was neutropenia (8%). AE’s of all grades occurring in >20% of patients were limited to diarrhea (24%), cough (22%), fatigue (20%), and nausea (20%). Notably, in comparison to other PI3Kδ inhibitors, no hepatotoxicity and no cases of colitis have been observed to date. Rates of infection and pneumonia have also been low (12% and 6%, respectively), and no cases of febrile neutropenia have been reported. Of the 41 patients treated at ≥ 800 mg of the previous formulation or with the micronized formulation, 32 are evaluable for efficacy (6 too early to evaluate, 2 non-compliant, 1 did not meet I/E criteria). Responses have been limited in patients with aggressive lymphoma and HL. Of the 9 evaluable CLL patients, 8 (89%) achieved a nodal PR (median nodal reduction of 71%), of which 5 achieved a PR per Hallek 2008 criteria with the remaining 4 having persistent lymphocytosis. The 1 CLL patient with SD had a >40% nodal reduction and remains on study. Of the 7 evaluable FL patients, all have shown clinical benefit with a reduction in tumor burden with 2 having achieved a PR, and the remaining 5 patients in SD. Additionally 2 MZL patients each achieved SD with >25% nodal reductions and remain on study. Notably, no patient with CLL or indolent lymphoma (FL & MZL) treated at ≥800 mg has progressed to date (median time on study of 20 weeks, range 6 – 73+), and no patient who achieved >50% reduction in tumor burden (including patients with CLL, FL, and HL) has progressed, with median time on study of 34 weeks (range 7 – 68+). Pharmacodynamic analysis in CLL patients indicates rapid suppression of pAKT at doses of 400 mg QD of the previous formulation. Conclusions: TGR-1202 is well tolerated in patients with relapsed and/or refractory hematologic malignancies with no reported hepatotoxicity or events of colitis and promising clinical activity. Enrollment continues in expansion cohorts and with the micronized formulation. Disclosures Brander: Celgene: Mentor received research funding Other. O'Connor:Celgene: Consultancy; Millennium Pharmaceuticals: Consultancy. Miskin:TG Therapeutics, Inc.: Employment, Equity Ownership. Sportelli:TG Therapeutics: Employment, Equity Ownership. Vakkalanka:Rhizen: Employment, Equity Ownership. Flinn:Infinity Pharmaceuticals: Consultancy.

Download Full-text

Comparison of Pharmacokinetics and Pharmacodynamics of Two Anti-CD20 Monoclonal Antibodies (Candidate Biosimilar DRL-Rituximab and Innovator Reference Product Rituximab (Mabthera®) in a Randomised, Multi-Centre, Double-Blind, Parallel Group Study of CHOP with Rituximab Chemotherapy in Patients with CD20-Positive Diffuse Large B-Cell Lymphoma

Blood ◽

10.1182/blood.v128.22.5391.5391 ◽

2016 ◽

Vol 128 (22) ◽

pp. 5391-5391

Author(s):

Auro Viswabandya ◽

Asis Mukhopadhyay ◽

Sandip Shah ◽

Rajnish Vasant Nagarkar ◽

Sonica Sachdeva Batra ◽

...

Keyword(s):

B Cell ◽

Cell Lymphoma ◽

Similarity Criterion ◽

B Cell Lymphoma ◽

Employment Equity ◽

Double Blind ◽

Large B Cell Lymphoma ◽

Parallel Group ◽

Equity Ownership ◽

Grade 3

Abstract Background: DRL-Rituximab (T) is a proposed biosimilar of rituximab. The similarity of DRL-Rituximab andtheinnovator reference products(USA: Rituxan®; European Union: Mabthera® [R]) has been demonstrated in physicochemical analyses and nonclinical studies. This study was undertaken to compare the pharmacokinetics (PK) and pharmacodynamics (PD) of T with R sourced from the European Union. Methods: Multiple-center, randomized, double-blind, two-arm, parallel-group study conducted in patients of both genders, aged 18-60 with a centrally confirmed newly diagnosed CD20 positive diffuse large-B-cell lymphoma (DLBCL) in their first line of treatment. Patients were randomized to receive 6 cycles of 21 days either T or R at an initial dose of 375 mg/m2 along with chemotherapy with cyclophosphamide, doxorubicin, vincristine and prednisone/prednisolone (CHOP). The primary objective of this study was to compare the PK of T and R. One of the secondary objectives was to establish the similarity of T and R in the pharmacodynamic parameters - as B-cell depletion and repletion in peripheral blood using counts of CD19+ positive cells determined by flow cytometry. The PK similarity criterion was that the 90% confidence intervals (CI) of the geometric mean ratios of AUC0-21days, and Cmax in Cycle 1 were within 80.00% - 125.00%. Cycle 1 PK, PD and summary safety results are reported here. Other PK parameters were evaluated as secondary endpoints. Results: A total of 151 patients were randomized to the study (76 to T, 75 to R). After exclusion of 2 patients with very low exposure (results confirmed to be outliers by a z-score higher than 3 and currently under further investigation), both products met the preset Cycle 1 PK similarity criterion (Table 1), with these 2 patients included the Cmax criterion was met, but for AUC the lower limit of the CI was 78.36% (Table 2). PK parameter values were generally comparable (Table 3). For PD both products appear comparable regarding both B-cell depletion and B-cell repletion, both in terms of proportion of patients as well as median time to depletion/repletion. (Table 4 & Figures 1A and 1B) Safety was also comparable between both products. Grade 3/4 Adverse Events (AEs) showed a similar trend in incidence across the two treatment arms [overall: 121 (80.1%); T: 57 (75.0%); R: 64 (85.3%)]. Neutropenia was the most frequently reported of the grade 3/4 AEs, followed by febrile neutropenia, leukopenia, and anaemia. The most commonly reported TEAEs related to the study drug were neutropenia [overall: 48 (31.8%); T: 29 (38.2%); R: 19 (25.3%)] and leukopenia [overall: 24 (15.9%); T: 16 (21.1%); R: 8 (10.7%)]. A total of 5 deaths are reported in the study amounting to 3.3% fatality rate. Conclusion: DRL-rituximab has equivalent PK profile to Mabthera in Cycle 1 when given in combination with standard CHOP chemotherapy for the treatment of CD20 positive DLBCL. PD and safety results are also similar. While awaiting results of further PK/PD data as well as outcomes from this trial, these results are highly encouraging and provide further impetus to development of DRL-rituximab as a candidate rituximab biosimilar. Acknowledgment: We thank all the Principal Investigators for their participation in conducting this study: Dr. Randeep Singh, Dr. Chiramana Haritha, Dr. Asis Mukhopadhyay, Dr. Anup Majumdar, Dr. Shailesh Bondarde, Dr. Rajnish Vasant Nagarkar, Dr. Vijay Ramanan, Dr. Chetan Dilip Deshmukh, Dr. MVT Krishna Mohan, Dr. SVSS Prasad, Dr. Nalini Kilara, Dr. Poonam Patil, Dr. Shekhar Patil, Dr. Neelesh Reddy, Dr. Suresh Sudalaiandi, Dr. Krishnan Srinivasan, Dr. Sundar Subramanian, Dr. VP Gangadharan, Dr. Auro Viswabandya, Dr. Alok Srivastava, Dr. Sharat Damodar, Dr. Rajesh Grover, Dr. Reena Nair, Dr. Bhausaheb Bagal, Dr. Kasi Viswanathan, Dr. Minish Jain, Dr. Jitendra Singh, Dr. Dipti Samanta, Dr. Sudha Sinha, Dr. Lakshmaiah Kuntegowdanahalli, Dr. Rajendersingh Arora, Dr. Sandip Shah, Dr. Shashikant Janardhan Apte, Dr. Mukul Goyal, and Dr. Prasanth Ganeshan. Disclosures Batra: Dr. Reddy's Labs: Employment, Equity Ownership. Lazaro:Dr. Reddy's Labs: Employment, Equity Ownership. Kankanwadi:Dr. Reddy's Labs: Employment, Equity Ownership.

Download Full-text

Blinatumomab Monotherapy Shows Efficacy in Patients with Relapsed Diffuse Large B Cell Lymphoma

Blood ◽

10.1182/blood.v118.21.1637.1637 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1637-1637 ◽

Cited By ~ 8

Author(s):

Andreas Viardot ◽

Mariele Goebeler ◽

Richard Noppeney ◽

Stefan W. Krause ◽

Stefan Kallert ◽

...

Keyword(s):

Adverse Events ◽

B Cell ◽

Phase Ii ◽

Research Funding ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Employment Equity ◽

Large B Cell Lymphoma ◽

Equity Ownership ◽

Large B Cell

Abstract Abstract 1637 Blinatumomab (MT103) is a single-chain bispecific antibody construct with specificity for CD19 and CD3 belonging to the class of bispecific T cell engager (BiTE®). A phase I trial with indolent and mantle cell lymphoma patients established a maximal tolerable dose (MTD) at 60 μg/m2/d. The trial was subsequently amended to evaluate blinatumomab in patients with diffuse large B cell lymphoma (DLBCL). Patients were treated by 4–8-week continuous i.v. administration with the following dosing regimen: first week at 5 μg/m2/d, second week at 15 μg/m2/d and for the remaining treatment period at 60 μg/m2/d. Two cohorts each with 6 DLBCL patients were enrolled. The two cohorts solely differed by the dose and schedule of corticosteroid medication administered at the beginning of blinatumomab infusion for mitigation of adverse events. In the first cohort 100 mg prednisolone was applied 1 hour prior to start; and in the second cohort patients received dexamethasone on days 1, 2, and 3. Three sequential patients received dexamethasone also 6–12 hours prior to start of infusion. Out of the twelve patients, 5 were male and 7 female. The median age was 57 years (range from 26 to 78 years). Patients had received a median of 4 prior regimens (range from 2–6). All patients had been exposed to rituximab. Eight of the 12 patients had undergone autologous stem cell transplantation (ASCT). International prognostic index (IPI) at screening ranged from 1 to 3 with a median of 2. The most common clinical adverse events (AEs) regardless of causality (>30%) were pyrexia (81.8%), fatigue (54.5%), constipation (36.4%), headache (36.4%), tremor (36.4%) and weight increase (36.4%). The most frequent laboratory AEs regardless of causality (>30%) were hyperglycemia (63.6%), lymphopenia (54.5%), C-reactive protein increase (45.5%), gamma-glutamyltransferase increase (45.5%) and thrombocytopenia (36.4%). Most AEs occurred early and were reversible. Four of 12 patients discontinued infusion due to fully reversible CNS events, 2 of which qualified as dose limiting toxicities (DLTs). Although just one DLT (reversible CNS event grade 3) occurred in the prednisolone cohort, a further cohort applying prophylactic dexamethasone was opened to optimize management of CNS events. A further refinement of the dexamethasone schedule, starting longer time prior to start of blinatumomab, was introduced after one early patient in the cohort receiving dexamethasone had experienced a reversible CNS event leading to discontinuation. All three patients treated in this manner completed the first blinatumomab cycle without discontinuations. Only one showed a grade 1 tremor, and no other CNS AEs were reported in these three patients. Two of 12 patients were not exposed to 60 μg/m2/d due to early discontinuations and 1 patient is too early in treatment for response evaluation. Five out of the remaining 9 evaluable patients (56%) showed objective clinical responses (4 CR/CRu; 1 PR). Three out of the 5 patients with CR/CRu or PR had prior ASCT. Two patients achieved objective responses (1 CR, 1 PR) despite of discontinuation at 60 μg/m2/d. The median response duration is +182 days (longest current duration +428 days), with 4 out of 5 responses still ongoing. Further evaluation of the last cohort will refine the recommended phase II dose, and the intensity and timing of dexamethasone comedication. The observation of lasting CRs after blinatumomab monotherapy in DLBCL patients is promising and warrants further exploration in a phase II study. Disclosures: Krause: Micromet: Research Funding. Mackensen:Micromet Inc.: Research Funding. Topp:Micromet: Consultancy, Honoraria. Scheele:Micromet Inc.: Employment, Equity Ownership, Patents & Royalties. Nagorsen:Micromet Inc.: Employment, Equity Ownership, Patents & Royalties. Zugmaier:Micromet: Employment. Degenhard:Micromet Inc: Employment. Schmidt:Micromet AG: Employment. Kufer:Micromet Inc: Employment, Equity Ownership. Libicher:Micromet Inc.: Consultancy, Honoraria. Bargou:Micromet: Consultancy, Honoraria.

Download Full-text

Differences In Treatment Patterns and Costs Among Diffuse Large B-Cell Lymphoma Patients Treated In The Clinic Vs. The Hospital Outpatient Setting

Blood ◽

10.1182/blood.v122.21.1751.1751 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1751-1751 ◽

Cited By ~ 1

Author(s):

Carolina Reyes ◽

Stacey Dacosta Byfield ◽

Laura K. Becker ◽

Art Small

Keyword(s):

Health Care ◽

B Cell ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Treatment Patterns ◽

Employment Equity ◽

Large B Cell Lymphoma ◽

Equity Ownership ◽

Large B Cell

Abstract Background Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of Non-Hodgkin's Lymphoma (NHL) accounting for approximately 30% of newly diagnosed casesi. DLBCL is an aggressive form of NHL and without treatment, median survival estimates are <1 year.ii Rituximab in combination with CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) is recommended first-line therapy for DLBCL patients and has been shown to improve overall survival compared with CHOP alone (previous standard therapy).iii In addition, published evidence suggests that receipt of granulocyte-colony stimulating factor (G-CSF) may improve outcomes among patients who initiate CHOP-based therapy.iv It is unclear whether differences in treatment and outcomes exist among cancer patients by site where care is delivered. This study examines differences in treatment patterns, health care resource use and costs among DLBCL patients receiving rituximab (R) or R+ chemotherapy in the office/clinic (OC) setting vs. the hospital outpatient (HOSP) setting. Methods This retrospective study used medical and pharmacy claims (1/2007 - 7/2012) from a national US commercial health plan to identify patients at least18 years old with ≥2claims for R. Patients were required to have evidence of DLBCL (≥1 claim with ICD-9-CM 200.78 or ≥2 claims with unique diagnosis codes from ICD-9-CM 200.70 to 200.77) and be enrolled in the health plan for ≥6 months before and after the index date (date of the first R claim). The follow-up period, that is, the episode of care (EOC), was the date of the first R infusion through 30 days after the last infusion prior to a gap in R administration of at least 7 months; those with less than 6 months of follow-up due to death were included. Patients with multiple cancers or receipt of R at both the OC and HOSP setting during the EOC were excluded. Differences in number of infusions, receipt G-CSF, healthcare utilization and per-patient per-month (PPPM) health care costs by cohort were examined. Results A total of 491 patients were identified, 65% OC (n=320) and 35% HOSP (n=171): by insurance type, 140 Medicare Advantage patients, 39% HOSP and 351 commercially insured patients, 33% HOSP. From 2007 to 2011/2012, the percentage of patients in HOSP increased from 32% to 43%. Descriptive results are shown in the Table. The cohorts had similar mean age, baseline Charlson comorbidity index scores and similar EOC lengths. However, compared to the OC cohort, the HOSP cohort had fewer infusions during the EOC and fewer infusions per month. In addition, fewer HOSP patients had evidence of combination therapy and receipt of any G-CSF during the EOC. HOSP patients also had significantly higher rates of emergency room visits, but not hospitalizations compared to OC patients. Total PPPM costs during the EOC as well as average costs of anti-cancer systemic therapy drugs plus administration costs incurred on days of rituximab infusions were significantly higher among the HOSP cohort compared to the OC cohort. Conclusions Increasing proportions of DLBCL patients receive infusions in the HOSP setting. HOSP patients had fewer infusions per month and incurred greater costs on the day of infusion compared to the OC cohort. There were fewer patients in HOSP with evidence of G-CSF during the EOC compared to OC patients. Overall, total PPPM costs were higher among the HOSP cohort compared to the OC cohort. Future research is warranted to assess the impact of these differences on clinical outcomes by site of care. [i] Armitage et al. JCO 1998;16(8):2780-95 [ii] Mey et al. Swiss Med Wkly 2012;140:w13511 [iii] NCCN Guidelines Version 1.2013 Diffuse Large B-cell Lymphoma [iv] Donnelly, et al, Leuk Lymphoma. 2000;39(1-2):67-75 Disclosures: Reyes: Genentech, inc: Employment, Equity Ownership. Dacosta Byfield:Genentech, Inc: Genentech contracted with OptumInsight to conducting the work described in the abstract. Stacey is employed at Optum but did not receive funds directly from Genentech and employment is not contingent on work with Genentech., Genentech contracted with OptumInsight to conducting the work described in the abstract. Stacey is employed at Optum but did not receive funds directly from Genentech and employment is not contingent on work with Genentech. Other; OptumInsight: Employment. Becker:Genentech, Inc: Genentech contracted with OptumInsight to conducting the work described in the abstract. Laura is employed at Optum but did not receive funds directly from Genentech and employment is not contingent on work with Genentech., Genentech contracted with OptumInsight to conducting the work described in the abstract. Laura is employed at Optum but did not receive funds directly from Genentech and employment is not contingent on work with Genentech. Other; OptumInsight: Employment. Small:Genentech, Inc: Employment, Equity Ownership.

Download Full-text

Mutation Impact of Targeted Genes in Diffuse Large B-Cell Lymphoma Patients Treated with Ibrutinib

Blood ◽

10.1182/blood.v126.23.2642.2642 ◽

2015 ◽

Vol 126 (23) ◽

pp. 2642-2642

Author(s):

Leo WK Cheung ◽

Karl J. Schweighofer ◽

Shiquan Wu ◽

Hsu-Ping Kuo ◽

Karl Eckert ◽

...

Keyword(s):

B Cell ◽

Clinical Response ◽

Cell Lymphoma ◽

Single Agent ◽

B Cell Lymphoma ◽

Good Clinical Response ◽

Employment Equity ◽

Mutation Impact ◽

Equity Ownership ◽

Tumor Biopsies

Abstract Introduction: Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin's lymphoma, accounting for approximately 30% of newly diagnosed cases in the United States. DLBCL can progress quickly and advanced cases are inconsistently cured with current therapies. Ibrutinib, a first-in-class Bruton's tyrosine kinase (BTK) inhibitor, is approved in the US and EU for patients with previously-treated mantle cell lymphoma and chronic lymphocytic leukemia (CLL) who have had one prior treatment, CLL with 17p deletion, and Waldenström macroglobulinemia. The activated B-cell (ABC) subtype of DLBCL is considered especially high risk and is characterized by chronic active B-cell receptor signaling, which is blocked by ibrutinib. Recent phase 2 trial results of ibrutinib as a single agent in DLBCL patients show an overall response rate of 41% in the ABC subtype (Wilson et al. ASH 2012; Wilson et al. Nat Med, 2015). Through targeted deep sequencing, we investigated the impact of baseline mutations of 317 targeted genes on clinical response of 51 DLBCL patients treated with ibrutinib. Based on our mutation impact analysis, we identified potential biomarkers for predicting DLBCL patient response to ibrutinib. In particular, we found sets of gene mutation patterns indicating poor (or good) clinical response across all subtypes (ABC, germinal-center B-cell-like [GCB], non-GCB) of DLBCL as well as uniquely within a subtype. Methods: H&E-stained slides from DLBCL patients enrolled in either PCYC-04753 (NCT00849654) or PCYC-1106 (NCT01325701) were reviewed to ensure sufficient nucleated cellularity and tumor content. DNA and RNA were extracted from unstained sections of FFPE DLBCL tumor biopsies. Sequencing was performed using the FoundationOne™ Heme panel following the validated NGS-based protocol to interrogate complete DNA coding sequences of 405 genes, as well as selected introns of 31 genes involved in rearrangements. RNA sequences of 265 commonly rearranged genes were analyzed to identify gene fusions. A subgroup of samples were analyzed on earlier versions of the FoundationOne™ panels where only DNA was extracted and sequenced. Sequence data was processed and analyzed for base substitutions, insertions, deletions, copy-number alterations, and selected gene fusions. Mutation impact indices (MII) of 317 genes were calculated and plotted for overall gene mutation pattern recognition. Chi-square association tests were performed on cases where sufficient sample sizes were available to determine statistical significance of mutation impact. DLBCL subtype classifications by gene expression profiling (GEP) and Hans' IHC were investigated and compared. For GEP, we used OmicSoft ArrayStudio's classification module to build linear discriminant analysis (LDA) model/classifier and neural networks with 5-fold cross validation procedure for model selection. The LDA was the best performing model and was selected for final GEP classification. Since only 29 (of 51) patients had central lab Hans' IHC classification information, we compared trends of the mutation impact results based on Hans' classification and GEP classification. Results: Single or multiple gene MII were generated from baseline tumor biopsies from DLBCL patients treated with single agent ibrutinib. The MII were generally consistent between GEP or Hans' IHC classification of tumor biopsies. We found novel baseline gene mutations associated with poor clinical response (SD or PD) to ibrutinib. These genes were involved in regulation of transcription (eg, mutations in EP300 in all DLBCL subtypes combined group [p=0.034], mutations in RB1 in ABC-DLBCL [p=0.031]), epigenetic modification (eg, mutations in MLL2 in ABC-DLBCL [p=0.053]), programmed cell death (mutations in BCL2 in all DLBCL subtypes [p=0.096]), and PI3K-AKT-mTOR pathway (eg, mutations in TSC2 in ABC-DLBCL [p=0.031]). Mutations identified as indicating good clinical response include mutations in CD79B (p=0.072) and MYD88 (p=0.024) in ABC-DLBCL. Co-existence of MYD88 and CD79B mutations (double-mutants) in ABC-DLBCL showed a stronger association to good clinical response (p=0.004) consistent with recent observations from Wilson et al. ASH 2012 & Nat Med, 2015. Conclusions: Our investigation reveals unique mutation patterns that underlie DLBCL subtypes and highlights the need for personalized medicine approaches to treating DLBCL patients. Disclosures Cheung: Pharmacyclics LLC, an AbbVie Company: Employment. Schweighofer:Pharmacyclics LLC, an AbbVie Company: Employment. Wu:Pharmacyclics LLC, an AbbVie Company: Employment. Kuo:Pharmacyclics LLC, an AbbVie Company: Employment. Eckert:Pharmacyclics LLC, an AbbVie Company: Employment. Balasubramanian:Pharmacyclics LLC, an AbbVie Company: Equity Ownership; Janssen: Employment, Equity Ownership. Ricci:Janssen: Employment, Equity Ownership, Patents & Royalties. Liang:Pharmacyclics LLC, an AbbVie Company: Employment. Beaupre:Pharmacyclics LLC, an AbbVie Company: Employment. Chang:Pharmacyclics LLC, an AbbVie Company: Employment.

Download Full-text

Lenalidomide Efficacy in Activated B-Cell-Like Subtype Diffuse Large B-Cell Lymphoma Is Dependent Upon IRF4 and Cereblon Expression

Blood ◽

10.1182/blood.v120.21.3287.3287 ◽

2012 ◽

Vol 120 (21) ◽

pp. 3287-3287

Author(s):

Ling-Hua Zhang ◽

Jolanta Kosek ◽

Maria Wang ◽

Carla Heise ◽

Peter H Schafer ◽

...

Keyword(s):

B Cell ◽

Cell Lines ◽

Molecular Mechanisms ◽

Cell Lymphoma ◽

Xenograft Model ◽

B Cell Lymphoma ◽

Cell Receptor ◽

Employment Equity ◽

Large B Cell Lymphoma ◽

Equity Ownership

Abstract Abstract 3287 Background: Durable responses with lenalidomide monotherapy have been reported in patients with non-Hodgkin lymphoma. In relapsed/refractory diffuse large B-cell lymphoma (DLBCL), higher responses were observed in the activated B-cell-like (ABC) subtype than in the germinal centre B-cell (GCB)-like subtype (Czuczman, et al. British Journal of Haematology, 2011, 154, 477–481). Herein, the molecular mechanisms involved in the differential efficacy of lenalidomide in DLBCL subtypes were investigated. Methods: A panel of DLBCL cell lines, with 5 of ABC-subtype and 11 of non-ABC subtype, was collected and cell of origin subtype was confirmed based on literature, molecular and genetic analysis. The direct antiproliferative effect of lenalidomide on DLBCL cells was assessed using the 3H-thymidine incorporation assay and apoptosis analysis. The molecular mechanisms involved in the antiproliferative efficacy of lenalidomide in DLBCL subtypes were investigated by western blot, immunohistochemistry (IHC) and qRT-PCR analysis of key signaling events during B-cell receptor (BCR)-dependent NF-κB activation. The critical roles of interferon regulatory factor 4 (IRF4), and cereblon (CRBN) in lenalidomide efficacy were established by knock-in or knock-down of these proteins in sensitive ABC cells. Finally, a mouse xenograft model was used to confirm the antitumor effect of lenalidomide and the relevance of the molecular mechanism involved. Results: Using DLBCL cell lines, lenalidomide treatment was found to preferentially suppress proliferation of ABC-DLBCL cells in vitro at a concentration range of 0.01–100 μM (the median plasma concentration at Cmax for patients receiving 25 mg lenalidomide is 2.2 μM) and delay tumor growth in a human tumor xenograft model of OCI-Ly10 cells (lenalidomide 3–30 mg/kg, p.o. qdX28), with minimal effect on non-ABC-DLBCL cells. This tumoricidal effect of lenalidomide was associated with downregulation of IRF4, a survival factor in ABC-DLBCL cells. Treatment with lenalidomide for 1–3 days, similar to the inhibitors of PKCb and MALT1 (LY-333,531 and z-VRPR-fmk, respectively), was found to significantly (p<0.05) downregulate IRF4 protein levels in sensitive cell lines such as OCI-Ly10 and U2932. IRF4 inhibition by lenalidomide reduced CARD11-BCL-10-MALT1 complex activity of ABC-DLBCL cells (as measured by BCL-10 cleavage) and resulted in downregulation of B-cell receptor (BCR)-dependent NF-κB activity. An NF-κB-driven luciferase assay revealed that lenalidomide (1 μM) inhibited transcriptional activity of NF-κB up to 56% in the sensitive ABC-DLBCL cell lines OCI-Ly10 (p <0.05) and U2932 (p <0.01) after 2-day drug treatment. Lenalidomide also significantly (p <0.05) inhibited DNA binding by Rel A/p65, p50 and c-rel/p70 in 4 lines of ABC cells. While IRF4-specific siRNA mimicked the effects of lenalidomide reducing NF-κB activation, IRF4 overexpression conferred cell resistance to lenalidomide, indicating the crucial role of IRF4 inhibition in lenalidomide efficacy in ABC DLBCL. Furthermore, knockdown of CRBN in OCI-Ly10 (p <0.05) and U2932 (p <0.01) conferred resistance to lenalidomide as demonstrated by the abrogation of the inhibitory effects of lenalidomide on IRF4 expression, BCL-10 cleavage, NF-κB activity, and proliferation of these cells, whereas the activity of inhibitors to PKC β and IKKα/β (LY-333,531 and CC-415501, respectively) remained unaffected. These data indicate that antitumor effects of lenalidomide on ABC-DLBCL cells require the presence of cereblon. Conclusions: These data may provide a mechanism for the preferential efficacy of lenalidomide in ABC-DLBCL observed in clinical studies. These findings suggest that lenalidomide has direct antitumor activity against DLBCL cells, preferentially ABC-DLBCL cells, by blocking IRF4 expression and the BCR-NF-κB signaling pathway in a cereblon-dependent manner (also see Figure below). Disclosures: Zhang: Celgene Corp: Employment, Equity Ownership. Kosek:Celgene Corp: Employment, Equity Ownership. Wang:Celgene Corporation: Employment, Equity Ownership. Heise:Celgene Corporation: Employment, Equity Ownership. Schafer:Celgene: Employment, Equity Ownership. Chopra:Celgene Corporation: Employment, Equity Ownership.

Download Full-text

Dual PI3Kδ/γ Inhibition By RP6530 Induces Apoptosis and Cytotoxicity In B-Lymphoma Cells

Blood ◽

10.1182/blood.v122.21.4411.4411 ◽

2013 ◽

Vol 122 (21) ◽

pp. 4411-4411 ◽

Cited By ~ 4

Author(s):

Swaroop Vakkalanka ◽

Srikant Viswanadha ◽

Eugenio Gaudio ◽

Emanuele Zucca ◽

Francesco Bertoni ◽

...

Keyword(s):

B Cell ◽

Cell Lines ◽

Whole Blood ◽

Cell Lymphoma ◽

Annexin V ◽

B Cell Lymphoma ◽

Primary Cells ◽

Employment Equity ◽

Human Whole Blood ◽

Equity Ownership

Introduction Activation of the PI3K pathway triggers multiple events including cell growth, cell cycle entry, cell survival and motility. While α and β isoforms are ubiquitous in their distribution, expression of δ and γ is restricted to cells of the hematopoietic system. Because these isoforms contribute to the development, maintenance, transformation, and proliferation of immune cells, dual targeting of PI3Kδ and γ represents a promising approach in the treatment of lymphomas. The objective of the experiments was to explore the therapeutic potential of RP6530, a novel, small molecule PI3Kδ/γ inhibitor, in B-cell lymphomas. Methods Activity and selectivity of RP6530 for PI3Kδ and γ isoforms and subsequent downstream activity was determined in enzyme and cell-based assays. Additionally, RP6530 was tested for potency in viability, apoptosis, and Akt phosphorylation assays using a range of immortalized B-cell lymphoma cell lines (Raji, TOLEDO, KG-1, JEKO, OCI-LY-1, OCI-LY-10, MAVER, and REC-1). Viability was assessed using the colorimetric MTT reagent after incubation of cells for 72 h. Inhibition of pAKT was estimated by Western Blotting and bands were quantified using ImageJ after normalization with Actin. Primary cells from lymphoid tumors [1 chronic lymphocytic leukemia (CLL), 2 diffuse large B-cell lymphomas (DLBCL), 2 mantle cell lymphoma (MCL), 1 splenic marginal zone lymphoma (SMZL), and 1 extranodal MZL (EMZL)] were isolated, incubated with 4 µM RP6530, and analyzed for apoptosis or cytotoxicity by Annexin V/PI staining. Results RP6530 demonstrated high potency against PI3Kδ (IC50=24.5 nM) and γ (IC50=33.2 nM) enzymes with selectivity over α (>300-fold) and β (>100-fold) isoforms. Cellular potency was confirmed in target-specific assays, namely anti-FcεR1-(EC50=37.8 nM) or fMLP (EC50=39.0 nM) induced CD63 expression in human whole blood basophils, LPS induced CD19+ cell proliferation in human whole blood (EC50=250 nM), and LPS induced CD45R+ cell proliferation in mouse whole blood (EC50=101 nM). RP6530 caused a dose-dependent inhibition (>50% @ 2-7 μM) in growth of immortalized (Raji, TOLEDO, KG-1, JEKO, REC-1) B-cell lymphoma cells. Effect was more pronounced in the DLBCL cell lines, OCI-LY-1 and OCI-LY-10 (>50% inhibition @ 0.1-0.7 μM), and the reduction in viability was accompanied by corresponding inhibition of pAKT with EC50 of 6 & 70 nM respectively. Treatment of patient-derived primary cells with 4 µM RP6530 caused an increase in cell death. Fold-increase in cytotoxicity as evident from PI+ staining was 1.6 for CLL, 1.1 for DLBCL, 1.2 for MCL, 2.2 for SMZL, and 2.3 for EMZL. Cells in early apotosis (Annexin V+/PI-) were not different between the DMSO blank and RP6530 samples. Conclusions RP6530 is a potent and selective dual PI3Kδ/γ inhibitor that inhibited growth of B-cell lymphoma cell lines with a concomitant reduction in the downstream biomarker, pAKT. Additionally, the compound showed cytotoxicity in a panel of lymphoma primary cells. Findings provide a rationale for future clinical trials in B-cell malignancies. Disclosures: Vakkalanka: Rhizen Pharmaceuticals, S.A.: Employment, Equity Ownership; Incozen Therapeutics Pvt. Ltd.: Employment, Equity Ownership. Viswanadha:Incozen Therapeutics Pvt. Ltd.: Employment. Bertoni:Rhizen Pharmaceuticals SA: Research Funding.

Download Full-text

Selective Inhibitor Of Nuclear Exports (SINE) KPT-330 In Combination With The Novel Proteasome Inhibitor Carfilzomib As a Novel Therapeutic Regimen For Treating Aggressive B-Cell Lymphomas

Blood ◽

10.1182/blood.v122.21.5147.5147 ◽

2013 ◽

Vol 122 (21) ◽

pp. 5147-5147

Author(s):

Elyse R Lopez ◽

Erin M Dominici ◽

Muychi T Vang ◽

Archito T. Tamayo ◽

Zhishuo Ou ◽

...

Keyword(s):

B Cell ◽

Cell Lines ◽

Nuclear Export ◽

Proteasome Inhibitor ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Selective Inhibitor ◽

The Novel ◽

Employment Equity ◽

Equity Ownership

Abstract Aggressive non-Hodgkin lymphomas (NHL), such as Diffuse Large B cell lymphoma (DLBCL) and mantle cell lymphoma (MCL), are one of a small number of important human cancers increasing in incidence in the US over the last four decades. Although these lymphomas are now potentially curable, almost half the treated patients still develop relapsed/refractory disease with poor survival outcomes, indicating an urgent need for better therapeutic approaches with improved efficacy. Double–hit lymphomas (DHL) constitute a particularly aggressive subtype of DLBCL and are characterized by a chromosomal breakpoint affecting the MYC/8q24 locus in combination with another recurrent breakpoint, usually a t(14;18)(q32;q21) involving BCL2. Prognosis for DHL is extremely poor, indicating a need for new treatment options as well. Multi-drug combination therapies are expected to potentiate therapeutic responses and delay disease recurrence by blocking adaptive resistance responses. Carfilzomib (CFZ), a novel second-generation proteasome inhibitor, and the novel Selective Inhibitor of Nuclear Exports (SINE) KPT-330 are effective treatments for refractory lymphomas. We hypothesized that combining the two would be even more successful because the drugs target different cellular pathways and processes in malignant B-cells. CFZ inhibits chymotrypsin-like, protein-degrading activity in the ubiquitin-proteasome pathway while KPT-330 irreversibly inhibits the major nuclear export receptor, chromosome region maintenance 1, CRM1, also termed XPO1. This study investigates whether the effects of CFZ and KPT-330 in combination are synergistic and thus more effective, reducing side effects and chemoresistance. Two DLBCL cell lines, DOHH-2 and OCI-Ly10, two DH-DLBCL cell lines, CJ and U-2973, and two MCL cell lines, Mino and Jeko-1, were used. Working with these cell lines, we identified and analyzed the increase in the efficacy of the combination over single agents alone through MTS proliferation and Annexin-V binding apoptosis assays. Growth inhibitory combination indexes for each cell line were calculated, and the Chou-Talalay Method was used to quantify additive effects (CI=1), synergistic effects (CI<1), or antagonistic affects (CI>1). Results indicated strong synergistic behavior of the combinatorial therapy, with many of the dose combinations indexes falling below 1. Apoptosis assays also showed a high degree of synergy when combining CFZ with KPT-330. The efficacy of CFZ and KPT-330 in combination signifies the combination’s potential as a foundation for future studies, treatments, and clinical trials. Disclosures: Kirk: Onyx Pharmaceuticals: Employment, Equity Ownership. Kauffman:Karyopharm Therapeutics Inc.: Employment. Shacham:Karyopharm : Employment, Equity Ownership, Membership on an entity’s Board of Directors or advisory committees, Patents & Royalties.

Download Full-text

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

Blood ◽

10.1182/blood.v126.23.2746.2746 ◽

2015 ◽

Vol 126 (23) ◽

pp. 2746-2746

Author(s):

Xiaoxian Zhao ◽

Darren C. Phillips ◽

Andrew J. Souers ◽

Juraj Bodo ◽

Paul Tapang ◽

...

Keyword(s):

Flow Cytometry ◽

B Cell ◽

Cell Lymphoma ◽

Ex Vivo ◽

B Cell Lymphoma ◽

Employment Equity ◽

Large B Cell Lymphoma ◽

Equity Ownership ◽

Cdk9 Inhibitor ◽

Large B Cell

Abstract 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. Disclosures Phillips: AbbVie: Employment, Other: Shareholder, Patents & Royalties. Souers:AbbVie: Employment, Equity Ownership. Tapang:AbbVie: Employment, Equity Ownership. Albert:AbbVie: Employment, Equity Ownership.

Download Full-text

The Dual Cell Cycle Kinase Inhibitor JNJ-7706621 Reverses Resistance to CD37 Targeted Radioimmunotherapy in Activated B Cell like Diffuse Large B Cell Lymphoma Cell Lines

Blood ◽

10.1182/blood-2019-123287 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 2574-2574

Author(s):

Gro Elise Rødland ◽

Katrine Melhus ◽

Roman Generalov ◽

Sania Gilani ◽

Francesco Bertoni ◽

...

Keyword(s):

B Cell ◽

Cell Lines ◽

Research Funding ◽

Kinase Inhibitor ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Employment Equity ◽

Large B Cell Lymphoma ◽

Equity Ownership ◽

Large B Cell

The CD37 targeting radioimmunoconjugate 177Lu-lilotomab satetraxetan (Betalutin®) is currently being evaluated as monotherapy in a clinical phase 2b trial for patients with follicular lymphoma (FL) and in a phase 1 trial for patients with diffuse large B-cell lymphoma (DLBCL), as well as in a phase 1b trial in combination with rituximab for patients with relapsed/refractory FL. Herein we have investigated the effect of 177Lu-lilotomab satetraxetan in seven activated B-cell like (ABC) DLBCL cell lines. Although the radioimmunoconjugate showed anti-tumor activity, primary resistance was observed in a subset of cell lines: U-2932 and RIVA. Both cell lines are representative for TP53 deficient Double Expressor (DE) DLBCL. Importantly, resistance was not a consequence of reduced binding of the radioimmunoconjugate to cell surface expressed CD37. Thus, we set out to identify drugs able to overcome the resistance to 177Lu-lilotomab satetraxetan in both resistant ABC-DLBCL cell lines. We performed a viability-based screen combining 177Lu-lilotomab satetraxetan with the 384-compound Cambridge Cancer Compound Library. Drug combinations were scored using Bliss and Chou-Talalay algorithms. We identified and characterized the dual-specific CDK1/2 and AURA/B kinase inhibitor JNJ-7706621 as compound able to revert the resistance to radioimmunotherapy (RIT), alongside topoisomerase and histone deacetylases (HDAC) inhibitors. Kinetic studies of the effect of mono- and combination therapy of U-2932 and RIVA cells with JNJ-7706621 and 177Lu-lilotomab satetraxetan are suggestive of a model in which radiation damage induced G2-arrested lymphoma cells eventually enter mitosis (repair or escape) and mitotic entry, progression and exit are impaired by JNJ-7706621 mediated inhibition of CDK1/2 and AURKA/B. Extended residence-time of cells in mitosis due to chromosome condensation and congression defects as well as spindle and mid-spindle assembly failure is likely pivotal for the increased sensitivity to persistent 177Lu-lilotomab satetraxetan deposited DNA damage, ultimately promoting cytokinesis failure (multinucleation, aneuploidy, increased cell size) and cell death. In conclusion, CD37-targeting 177Lu-lilotomab satetraxetan RIT showed activity in several ABC-DLBCL lymphoma cell lines. CD37-independent RIT-resistance was identified in two cell lines representative of aggressive DE ABC-DLBCLs with inactive TP53, and reversed by subsequent inhibition of CDK1/2 and AURKA/B by JNJ-7706621. These findings may be of potential relevance for ongoing clinical trials of 177Lu-lilotomab satetraxetan in relapsed, ASCT-non-eligible DLBCL, and may also be more generally applicable to other 177Lu-based RITs and alternative radionuclide utilizing targeted therapies. Future pre-clinical investigations are required to elucidate the potential application of CDK1/2 and AURKA/B inhibitors as a strategy to revert RIT resistance in TP53 deficient cancers. Disclosures Rødland: Nordic Nanovector ASA: Patents & Royalties, Research Funding. Melhus:Nordic Nanovector ASA: Employment, Equity Ownership, Patents & Royalties. Generalov:Nordic Nanovector ASA: Employment, Equity Ownership, Patents & Royalties. Bertoni:Nordic Nanovector ASA: Research Funding; Oncology Therapeutic Development: Research Funding; PIQUR Therapeutics AG: Other: travel grant, Research Funding; HTG: Other: Expert Statements ; Amgen: Other: travel grants; Astra Zeneca: Other: travel grants; Jazz Pharmaceuticals: Other: travel grants; NEOMED Therapeutics 1: Research Funding; Acerta: Research Funding; ADC Therapeutics: Research Funding; Bayer AG: Research Funding; Cellestia: Research Funding; CTI Life Sciences: Research Funding; EMD Serono: Research Funding; Helsinn: Consultancy, Research Funding; ImmunoGen: Research Funding; Menarini Ricerche: Consultancy, Research Funding. Dahle:Nordic Nanovector ASA: Employment, Equity Ownership, Patents & Royalties. Syljuåsen:Nordic Nanovector ASA: Patents & Royalties, Research Funding. Patzke:Nordic Nanovector ASA: Employment, Patents & Royalties.

Download Full-text