Blood

Nucleoporin 98 (NUP98) fusion oncoproteins are observed in a spectrum of hematologic malignancies, particularly pediatric leukemias with poor patient outcomes. Although wild-type full-length NUP98 is a member of the nuclear pore complex, the chromosomal translocations leading to NUP98 gene fusions involve the intrinsically disordered and N-terminal region of NUP98 with over 30 partner genes. Fusion partners include several genes bearing homeodomains or having known roles in transcriptional or epigenetic regulation. Based on data in both experimental models and patient samples, NUP98 fusion oncoprotein–driven leukemogenesis is mediated by changes in chromatin structure and gene expression. Multiple cofactors associate with NUP98 fusion oncoproteins to mediate transcriptional changes possibly via phase separation, in a manner likely dependent on the fusion partner. NUP98 gene fusions co-occur with a set of additional mutations, including FLT3–internal tandem duplication and other events contributing to increased proliferation. To improve the currently dire outcomes for patients with NUP98-rearranged malignancies, therapeutic strategies have been considered that target transcriptional and epigenetic machinery, cooperating alterations, and signaling or cell-cycle pathways. With the development of more faithful experimental systems and continued study, we anticipate great strides in our understanding of the molecular mechanisms and therapeutic vulnerabilities at play in NUP98-rearranged models. Taken together, these studies should lead to improved clinical outcomes for NUP98-rearranged leukemia.

Interleukin-3 (IL-3) is exclusively expressed by activated T and natural killer cells, a function that is tightly controlled both in a lineage-specific and in a stimulation-dependent manner. We have investigated the protein binding characteristics and functional importance of the ACT-1-activating region of the IL-3 promoter. This region binds an inducible, T-cell-specific factor over its 5′ end, a site that is necessary for the expression of IL-3 in the absence of other upstream elements. Over its 3′ end, it binds a factor that is ubiquitously and constitutively expressed. This factor is Oct-1 or an immunologically related octamer-binding protein, and it plays a role in coordinating the activity of several regulatory elements. These characteristics make the ACT-1 site analogous to the activating ARRE-1 site in the IL-2 promoter. Furthermore, and despite a lack of sequence homology, the promoters of IL-3 and IL-2 share an organizational pattern of regulatory elements that is likely to be important for the T- cell-specific expression of these genes.

Abstract 2476

Bendamustine is a unique alkylating agent which combines a nitrogen mustard moiety of mechlorethamine with a benzimidazole. This study was conducted to characterize the distribution, metabolism, and elimination of [14C] bendamustine and its metabolites (M3, M4, and dihydroxy bendamustine [HP2]) and to assess the roles of renal and hepatic pathways in the drug's metabolism and excretion. A secondary objective was to further characterize the safety profile of single-agent bendamustine.

This open-label, phase I study enrolled 6 patients, age ≥18 years, with confirmed relapsed or refractory malignancy. The study was divided into 2 assessment periods: period A, during which the mass balance and pharmacokinetics of [14C] bendamustine were investigated, and period B, an extended-use period of up to 6 cycles with non-labeled bendamustine, during which safety continued to be assessed. Patients received intravenous (IV) bendamustine (120 mg/m2), containing 80–95 μCi of [14C] bendamustine, on day 1 of cycle 1 and non-labeled IV bendamustine (120 mg/m2) on day 2 of cycle 1 (period A). Pharmacokinetic parameters of bendamustine and metabolites M3, M4, and HP2 were calculated through plasma and urine concentrations, which were determined through 24 hours following administration of bendamustine on day 1. Total radioactivity (TRA) levels were measured in plasma, urine, and feces collected prior to drug administration and at time points through 168 hours after patients received [14C] bendamustine. Collection of excreta could continue (after the 7-day period) on an outpatient basis: if radiolabeled bendamustine ≥1% of dose was measurable in the 144- to 168-hour urine or feces collection, collection continued until the recovery in each 24-hour urine or feces collection was <1% of dose.

Six patients (3 males; 3 females) with a median age of 66 (48–75) years were enrolled and completed the pharmacokinetic portion of the study. For bendamustine, the decline from peak plasma concentration was characterized by an initial rapid distribution phase, followed by a somewhat slower intermediate phase. The pharmacologically relevant half-life (t½) was approximately 40 minutes. The plasma concentrations of M3, M4, and HP2 were very low relative to the bendamustine concentrations.

Of the TRA dose administered, approximately half of the dose was recovered in the urine and approximately a quarter of the dose was recovered in the feces. Less than 5% of TRA dose was recovered in the urine as unchanged bendamustine. Mean recovery of TRA in excreta was approximately 76% of the radiochemical dose. Total recovery was incomplete due to continued slow excretion of TRA at the end of the collection period. The sustained levels of radioactivity in the plasma as compared with plasma concentrations of bendamustine suggest that, despite the rapid clearance of bendamustine, 1 or more longer-lived [14C] bendamustine-derived materials remain in the plasma. These longer-lived materials likely include by-products of alkylation. As previously noted, bendamustine volume of distribution was small (Vss∼20 L). The steady-state volume of distribution for TRA was ∼50 L. These results confirm previous data and provide evidence that neither bendamustine nor TRA are extensively distributed into the tissues. All 6 patients withdrew prior to completion of period B due to disease progression (n = 4), an adverse event (n = 1), or refusal to continue treatment (n=1). Bendamustine was well tolerated when administered at a dosage of 120 mg/m2 for 2 to 3 cycles. The most frequent treatment-related adverse events were fatigue (50%) and vomiting (50%). A grade 3/4 absolute lymphocyte count decrease occurred in all patients at some point during the study. There were no other grade 3/4 hematologic adverse events.

Bendamustine was extensively metabolized via multiple metabolic pathways, with subsequent excretion in both urine and feces. Bendamustine accumulation is not anticipated in cancer patients with renal or hepatic impairment due to the dose administration schedule and short intermediate half-life. Adverse events and hematologic changes were consistent with the known safety profile of bendamustine.

This research was sponsored by and conducted by Cephalon, Inc., Frazer, PA.

Darwish: Cephalon, Inc.: Employment. D'Andrea:Cephalon, Inc.: Employment. Bond:Cephalon, Inc.: Employment. Hellriegel:Cephalon, Inc.: Employment.

Fusion genes derived from the platelet-derived growth factor receptor beta (PDGFRB) or alpha (PDGFRA) play an important role in the pathogenesis of BCR-ABL–negative chronic myeloproliferative disorders (CMPDs). These fusion genes encode constitutively activated receptor tyrosine kinases that can be inhibited by imatinib. Twelve patients with BCR-ABL–negative CMPDs and reciprocal translocations involving PDGFRB received imatinib for a median of 47 months (range, 0.1-60 months). Eleven had prompt responses with normalization of peripheral-blood cell counts and disappearance of eosinophilia; 10 had complete resolution of cytogenetic abnormalities and decrease or disappearance of fusion transcripts as measured by reverse transcriptase–polymerase chain reaction (RT-PCR). Updates were sought from 8 further patients previously described in the literature; prompt responses were described in 7 and persist in 6. Our data show that durable hematologic and cytogenetic responses are achieved with imatinib in patients with PDGFRB fusion–positive, BCR-ABL–negative CMPDs.

Introduction: Central nervous system (CNS) relapse is an uncommon but devastating complication occurring in about 2-10% of patients with diffuse large B-cell lymphoma (DLBCL). The CNS-IPI is a validated prognostic tool used to identify patients at high risk of CNS relapse. MYC and BCL2/BCL6 rearrangements (double-hit lymphoma- DHL) and non-GCB (NGCB) cell of origin may be at higher risk of CNS relapse. However, the ability of these molecular data to refine CNS-IPI risk prediction has not been assessed. In this study, we aimed to describe the impact of DHL and NGCB cell of origin in the incidence of CNS relapse in patients with newly diagnosed DLBCL.

Methods: A retrospective review was conducted to identify patients with newly diagnosed DLBCL from 01/01/2002 - 06/30/2015 using the University of Iowa/Mayo Clinic Lymphoma Molecular Epidemiology Resource database. Patients with primary CNS lymphoma or with CNS involvement at diagnosis were excluded. All patients received chemoimmunotherapy. Clinical, biological, laboratory and radiological data were abstracted including the occurrence of CNS relapse. CNS IPI risk score categories were defined as low- (≤1) intermediate- (2-3) and high-risk (≥4) as originally described (Schmitz et al, JCO 2016). Cell of origin (COO) classification into GCB and non-GCB subtypes was performed using Hans algorithm and/or NanoString data. Characteristics between groups were compared using Kruskal-Wallis or Chi-squared test as appropriate. Survival analysis was performed using Cox proportional hazards model and the Kaplan Meier method. Statistical analysis was performed using SAS version 9.4M5 and R version 4.0.3.

Results: We identified a total of 1,360 patients with newly diagnosed DLBCL. Patients had a median age of 62 years (range 18-93 years), were predominantly male (56%), had a performance status &lt;2 (84%), had stage III/IV disease (61%) and an elevated LDH (56%) at diagnosis. Fluorescent in situ hybridization data was present for 65% of patients. Among patients with available data, DHL and NGCG COO represented 2.8% and 36% of the cohort, respectively. CNS IPI high- and intermediate-risk categories were present in 12.4% and 52% of the patients, respectively. R-CHOP was the most common upfront chemotherapy (n=980, 72%).

The median follow-up for the entire cohort was 84.9 months (IQR: 58.9-120.9). Overall, 50 patients (3.6%) had a CNS relapse during the study period. Lymphoma relapsed in the CNS in 3.0% (95% CI:2.1-4.4), 4.7% (95% CI: 3.4-6.3) and 4.9% (95% CI: 3.7-6.6)% of patients with CNS-IPI score of intermediate/high risk at 1, 3 and 5 years following the diagnosis of DLBCL. In patients with low risk CNS-IPI, 1.1% (95% CI: 0.4-2.5), 1.7% (95% CI: 0.9-3.4) and 1.7% (95% CI: 0.9-3.4) had a CNS relapse at 1, 3 and 5 years following DLBC diagnosis. Patients with CNS relapse were more likely to have elevated LDH (76% vs 56%, p=0.008), advanced stage (78% vs 61%, p=0.01), adrenal involvement (8% vs 2.4%, p=0.014) and have a CNS IPI intermediate risk (68% vs 51%), and high risk (16% vs 12.3%, p=0.014) at the time of initial diagnosis compared to patients without CNS relapse. Patients with CNS relapse had similar rates of DHL (4% vs 2.7%, p=0.17) and NGCB cell of origin (42% vs 36.2%, p=0.52) compared to patients without CNS relapse.

In the multivariable analysis, the strongest predictors for CNS relapse were CNS IPI high risk (Hazard Ratio [HR]= 3.6, 95%CI= 1.4-9.7) and CNS IPI intermediate risk (HR= 3.15, 95%CI= 1.46-6.81). DHL and NGCB cell of origin were not associated with a significantly higher risk of CNS relapse (HR= 1.79, [95%CI=0.42-7.59] and HR= 1.26, [95%CI=0.62-2.57], respectively).

Conclusion: CNS relapse was an uncommon occurrence in patients with DLBCL who were treated with chemoimmunotherapy. CNS IPI intermediate- and high-risk scores remain the strongest predictors of CNS relapse in patients with DLBCL after adjusting to biological high-risk variables including DHL and NGCB cell of origin. Larger studies are required to validate our findings due to the low incidence of CNS relapse in our cohort.

Figure 1 Figure 1.

Farooq: Kite, a Gilead Company: Honoraria. Maurer: Kite Pharma: Membership on an entity's Board of Directors or advisory committees; Morphosys: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Research Funding; Genentech: Research Funding; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees; Nanostring: Research Funding. Witzig: Karyopharm Therapeutics, Celgene/BMS, Incyte, Epizyme: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene/BMS, Acerta Pharma, Kura Oncology, Acrotech Biopharma, Karyopharm Therapeutics: Research Funding. Habermann: Tess Therapeutics: Other: Data Monitoring Committee; Seagen: Other: Data Monitoring Committee; Incyte: Other: Scientific Advisory Board; Morphosys: Other: Scientific Advisory Board; Loxo Oncology: Other: Scientific Advisory Board; Eli Lilly & Co.,: Other: Scientific Advisor. Novak: Celgene/BMS: Research Funding.

Introduction

Cold agglutinin disease (CAD) is a serious and rare autoimmune hemolytic anemia driven by cold agglutinin autoantibodies, which bind to red blood cells and activate the classical complement system to initiate hemolysis and anemia (Berentsen S. Hematology Am Soc Hematol Educ Program 2016). There is limited evidence on the individual and societal impact of CAD. A retrospective study of 27 patients in a US healthcare institute demonstrated fluctuations in severity of anemia over the course of the disease, and significant utilization of healthcare resources (Mullins M et al. Blood Adv 2017). The objective of this study was to understand the long-term characteristics and disease burden in patients with primary CAD from a large US Electronic Health Record (EHR) database.

Methods

This retrospective observational cohort study included adult patients from the Optum© EHR database between January 1, 2007 and September 30, 2019 who had ≥1 medical encounter with an autoimmune hemolytic anemia-related diagnosis code, ≥3 documentations (on different dates) of CAD and ≥1 hemoglobin (Hb) value &lt;12 g/dL. The index date was defined as the first mention of CAD; all patients were required to have a 12-month baseline period prior to this. To limit the study to patients with primary CAD, patients were excluded if they had ≥1 medical encounter with mycoplasma, cytomegalovirus and Epstein-Barr at the index date, or ≥1 medical encounter with lymphoma, MALT lymphoma, chronic lymphoid leukemia, Waldenstrom macroglobulinemia or myeloma during the baseline period.

Anemia severity (defined as the lowest Hb value in each study period), utilization of CAD-related therapies, blood transfusions and all-cause healthcare resource utilization (HCRU) were analyzed at baseline and at 6-month follow-up intervals. Although no treatment is approved in CAD, corticosteroids, immunoglobulin, rituximab, immunosuppressants, antineoplastic and biologics were considered as CAD-related therapies to reflect usual practice. Results were stratified by anemia severity category (severe [Hb &lt;8 g/dL], moderate [Hb 8.0-10 g/dL], mild [Hb 10.1-&lt;12 g/dL] or no anemia [Hb ≥12 g/dL]) during each follow-up interval. Severe hemolysis was defined as elevated LDH and/or elevated bilirubin.

Results

A total of 610 adults with primary CAD were included in the study (mean [SD] age 67.9 [14.5] years, 65.4% female). The mean (SD) duration of follow-up was 48.1 (30.6) months; 90% of patients had ≥12 months of follow-up. At baseline (0-6 months prior to first mention of CAD), 47.6% of patients had elevated bilirubin levels and 63.1% had elevated lactate dehydrogenase (LDH) levels.

A high proportion of patients with CAD experienced severe or moderate anemia at baseline and in the 6 months post-baseline; this proportion tended to be lower, but still substantial, throughout the follow-up period (Table). Frequency of moderate/severe anemia or severe hemolysis events per patient year was also higher in the first 6 months: 5.70 (95% CI: 5.00, 6.49), compared with 2.92 (2.30, 3.71) and 2.43 (1.89, 3.11) events at months 19-24 and months 31-36, respectively.

The median number of CAD-related treatments per patient was high in all CAD patients at 6 months and remained high during the follow-up period (Table). The most common therapies used (excluding blood transfusion) were corticosteroids, antineoplastics and biologics. The mean number of blood transfusions per patient was higher in the severe anemia category at all follow-up intervals. The number of hospitalizations and emergency room visits were generally higher in patients with increased anemia severity; outpatient visits were high in all CAD patients and remained so over the study period (Table).

Conclusion

This observational cohort study followed a large sample of primary CAD patients with a 4-year mean follow-up. The results highlight the long-term substantial burden of CAD on patients and healthcare systems, which generally increased with higher severity of anemia. Three years after diagnosis, the number of moderate to severe anemia or hemolysis events remained high in CAD patients, despite off-label CAD management. The need for blood transfusions was still substantial in the severe anemic population 3 years after diagnosis. This longitudinal analysis illustrates the unmet medical needs in primary CAD.

Wilson: Sanofi: Current Employment, Current equity holder in publicly-traded company. Joly:Sanofi: Current Employment, Current equity holder in publicly-traded company. Miles:Aetion Inc.: Current Employment, Other: Employee of software company that received funding from Sanofi for the current study. Kuang:Aetion Inc.: Current Employment, Other: Employee of software company that received funding from Sanofi for the current study. Pham:Sanofi: Consultancy; Alexion: Other: Speaker.