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Background: Classical Hodgkin lymphoma (HL) has a unique histopathology, with rare malignant Hodgkin/Reed Sternberg (HRS) cells surrounded by a strong inflammatory cellular component in the tumor microenvironment (TME). Although extensive studies describe the interdependence of the HRS cells and the TME, the impact of HL on systemic immunity has not been well described. Here, we develop a new approach, employing a recently commercialized single cell cytokine secretion platform (IsoLight) to assess, precisely and comprehensively, the function of peripheral blood mononuclear cells (PBMCs) in HL patients. Methods: Cells were selected from 4 HL patients: 2 newly diagnosed who had a complete response (CR) to therapy standard first line therapy, and 2 relapsed patients who progressed on second line chemotherapy (PD). Cryopreserved PBMCs from a pre-treatment and post-treatment time point for each patient were thawed, rested overnight, stimulated with PMA/ionomycin and loaded into the IsoLight single cell cytokine secretion system. IsoLight captures single cells in microwells; as cytokines are secreted, they are bound by antibodies lining the microwell cover. Bound cytokines are then revealed by fluorescent secondary antibodies and photos are taken at various time points to assess fluorescence intensity, which corresponds to the relative amount of each cytokine secreted. Twenty thousand cells can be assayed per sample simultaneously. Results: The percentage of cytokine-secreting cells varied dramatically by donor (12%-48%), with monofunctional cells making only TNFa, MIP1b, or IL-15 dominating the functional landscape. Polyfunctional cells, capable of making three or more cytokines simultaneously represented only 0.1-7% of the cells in each sample, but there were more of these cells, and each secreted higher levels of cytokines, in individuals who responded to therapy with a CR. Responders also secreted higher levels of IL2, Perforin, IL4, IL12, MIP1a, and TNFb (p values ranging from 0.005 to 0.03), and lower levels of IL9 and IL22 (p=0.0028 and 0.021, respectively), compared to non-responders at diagnosis. Responders lost expression of IL4, IL7, and MIP1a over the course of treatment (pre- vs post-treatment, p=0.01 to 0.05), while non-responders gained cells that expressed IL4, IL5, IL10, IL17, and TNFb from diagnosis to end of treatment (p=0.001 to 0.05). Conclusion: This work represents an important methodological advance in immune monitoring for hematologic malignancies. Single cell cytokine secretion technology measures more cytokines simultaneously than flow cytometry, providing a sample-sparing and comprehensive overview of the functional landscape of immune cells in a patient. Moreover, the technology provides cell-by-cell information about cytokine secretion, unlike Luminex. Our work represents the first application of this technology to HL, which we use to define, for the first time, the particular combinations of 32 cytokines that can be secreted by individual immune cells. We also identify candidate cytokines whose frequency at diagnosis may predict treatment outcome, and reveal changes in cytokine levels over treatment time that may distinguish patients destined to relapse. Immunotherapy may impact PBMC function differently, this may partially explain the high efficacy of this therapy in the relapsed population. The impact of immunotherapy on cytokine levels is currently under investigation by our group in a larger study. Other important questions which are under investigation include the impact of prior chemotherapy on cytokine profiles in relapsed patients, and whether certain cytokines which increase during treatment may be a surrogage for tumor bulk in patients with PD. Cytokines elevated in patients with poor responses to treatment include IL9, IL10, IL17, and IL22, which may present attractive drug targets if validated in our larger ongoing follow-up study. Disclosures Diefenbach: Bristol-Myers Squibb: Consultancy, Research Funding; MEI: Research Funding; Denovo: Research Funding; Genentech: Consultancy, Research Funding; Incyte: Research Funding; LAM Therapeutics: Research Funding; Merck: Consultancy, Research Funding; Seattle Genetics: Consultancy, Research Funding; Trillium: Research Funding; Millenium/Takeda: Research Funding. Hymes:Celgene: Consultancy. Martin:Janssen: Consultancy; Sandoz: Consultancy; Karyopharm: Consultancy; Celgene: Consultancy; Teneobio: Consultancy; I-MAB: Consultancy. Ruan:Celgene: Consultancy, Honoraria, Research Funding; AstraZeneca: Consultancy, Honoraria; Seattle Genetics: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria; Pharmacyclics LLC, an AbbVie company: Research Funding; Juno: Consultancy; Kite: Consultancy. Leonard:Miltenyi: Consultancy; Akcea Therapeutics: Consultancy; Sandoz: Consultancy; AstraZeneca: Consultancy; Bayer Corporation: Consultancy; Epizyme, Inc: Consultancy; BeiGene: Consultancy; Miltenyi: Consultancy; ADC Therapeutics: Consultancy; Akcea Therapeutics: Consultancy; Sandoz: Consultancy; Celgene: Consultancy; Epizyme, Inc: Consultancy; Karyopharm Therapeutics: Consultancy; AstraZeneca: Consultancy; Bayer Corporation: Consultancy; Celgene: Consultancy; Sutro Biopharma: Consultancy; Merck: Consultancy; Genentech, Inc./F. Hoffmann-La Roche Ltd: Consultancy; Gilead: Consultancy; Karyopharm Therapeutics: Consultancy; Sutro Biopharma: Consultancy; Nordic Nanovector: Consultancy; ADC Therapeutics: Consultancy; MorphoSys: Consultancy; Gilead: Consultancy; Nordic Nanovector: Consultancy; BeiGene: Consultancy; Merck: Consultancy; Genentech, Inc./F. Hoffmann-La Roche Ltd: Consultancy; MorphoSys: Consultancy.

Background: Over-expression of Aurora A has been shown in many subtypes of non-Hodgkin lymphoma (NHL) and is a negative prognostic factor in mantle cell lymphoma (MCL), where expression correlates with tumor proliferative signature (Ki67), and may contribute to cell cycle dysregulation. Aurora A regulates chromosomal stability through effects on mitosis and cell cycle regulation. Bortezomib increases both pro-apoptotic proteins and cell cycle dependent kinase inhibitors. Thus, there is a strong scientific rationale to combine Aurora A kinase and proteasome inhibition. We evaluated the combination of alistertib, an oral Aurora A kinase inhibitor, with bortezomib and rituximab in a CTEP sponsored phase 1 study of patients with relapsed /refractory (RR) low grade NHL or MCL. Methods: Patients with RR low grade NHL or MCL, who had received at least one line of immuno-chemotherapy were treated according to a 3 + 3 design with 3 dose escalation cohorts (DL1, DL2, and DL3) to identify the recommended phase 2 dose (RP2D) with an expansion cohort at the RPTD. Patients received alisertib 30mg BID (DL1 and DL2) or 40mg BID (DL3) on days 1-7, bortezomib subcutaneous 1mg/m2 (DL1) or 1.3 mg/m2 (DL2 and DL3) on days 1, 8, and 15, and rituximab 375mg/m2 on day 1 for the first 8 cycles, and subsequently q 3 months. An expansion cohort was treated at DL3. Treatment cycles were 28 days. Dose Limiting Toxicity (DLT) was defined within the first cycle of therapy. Results: A total of 24 patients at 3 sites were enrolled between 06/2013 and 02/2018, including DL1: 3, DL2: 7; DL3: 14. Median age was 64 (range 46-87). Twelve patients (50%) were male. Nineteen patients had low grade NHL, and 5 patients had MCL. Performance status was 0-1 in 22 (92%) patients, and 2 in 2 (8%) patients. Dose Escalation and Safety: The recommended phase II dose (RP2D) was DL3, with the only DLT in DL3 (prolonged grade 4 neutropenia). The most common grade 3-4 AEs at all dose levels were neutropenia (34%), thrombocytopenia (13%), anemia (8%), and fatigue (8%); grade 2 alopecia occurred in 37% (Table 1). Other grade 4 AEs included hypertension & hypotension, respiratory failure, hyperkalemia and hyperuricemia (n=1 (4%) each); one patient died of GI hemorrhage deemed unrelated to study treatment (NSAID overdose). Grade 3 neutropenia and leukopenia was seen in 4 patients and thrombocytopenia in 1. There was one grade 3 AE each of: hypertension, pulmonary hypertension, pneumonitis, fatigue, diarrhea, infusion reaction, lung infection, and febrile neutropenia. Efficacy: A total of 18 patients were evaluable for response. Five patients were not evaluable for response for the following reasons: withdrawal of consent (3, 1 each DL1, 2, 3), non-compliance (1 DL2), and death unrelated to study drug (1, DL2). For patients who were treated with at least 3 cycles of therapy, and had a restaging CT scan, the median duration of therapy was 142 days, (range 29-590). Response was seen in 7/18 evaluable patients for an overall response rate (ORR) of 38%; 4 of 18 patients (22%) had a complete response (CR), and 4 of 18 patients (22%) had a partial response (PR). An additional 8/18 patients (44%) had stable disease for a clinical benefit rate of 88%. With a median follow-up of 30.9 months, the median PFS was 6.9 months (95%CI 4.1-28.0) and median OS not reached. OS at 3 years was 64.9% (95% CI 44.1-95.3%; Fig. 1). Median duration of response was 14.1 mo (95% CI 2.63-NR). Conclusion: Alisertib in combination with bortezomib and rituximab is a well-tolerated regimen with significant and durable activity in heavily pretreated low grade NHL and MCL. The RP2D is DL3 (R + bortezomib 1.3mg/m2 and 1.8 alisertib 40mg bid). A correlation of Aurora Kinase tumor expression with clinical outcome is planned. Disclosures Barta: Janssen: Membership on an entity's Board of Directors or advisory committees; Merck, Takeda, Celgene, Seattle Genetics, Bayer: Research Funding. Diefenbach:Merck: Consultancy, Research Funding; Genentech: Consultancy; Incyte: Research Funding; Millenium/Takeda: Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Acerta: Research Funding; Seattle Genetics: Consultancy, Research Funding; Trillium: Research Funding; Denovo: Research Funding.

Background: The CALGB 10603 RATIFY trial demonstrated that midostaurin, a multi-targeted small molecule FLT3 inhibitor, when given in combination with standard 3+7 induction consisting of daunorubicin 60mg/m2 for 3 doses and cytarabine 200mg/m2/day for 7 days significantly prolonged overall survival (OS) compared to placebo plus standard induction in FLT3 positive acute myeloid leukemia (AML) patients. The safety and efficacy of the other anthracycline commonly utilized in standard AML induction, idarubicin, has not been evaluated to date. Methods: A single-center retrospective review from May 2017 to July 2018 was performed. Patients were included if they had a diagnosis of FLT3 positive AML and received induction chemotherapy with idarubicin. The primary outcome was incidence of adverse effects attributed to midostaurin. Grading of adverse effects was in accordance with the Common Terminology and Criteria of Adverse Effects (CTCAE) version 5. Additional outcomes included OS, relapse-free survival (RFS), similar as in the Ratify trial, as well as detection of FLT3 on subsequent bone marrow biopsies. Results: Ten patients were included. All patients received induction therapy with idarubicin 12mg/m2 for 3 doses and cytarabine 100mg/m2/day for 7 days. Median age was 53 years (range: 33 to 66) and 6/10 were male. Eight of 10 patients exhibited internal tandem duplication (ITD) on diagnosis; two had FLT3 tyrosine kinase domain (TKD) D835. Eight patients had diploid cytogenetics; the other two patients had core-binding factor AML. Midostaurin was initiated on day 8 of induction in all but 2 patients, who started on day 11 and 12, respectively. Nine of ten patients completed all 28 planned doses of midostaurin. All patients received antifungal prophylaxis with micafungin throughout the course of midostaurin. The median time from day 1 of induction to neutrophil (>500/µl) and platelet (>100,000/µl) recovery was 23 days and 26 days, respectively. Granulocyte colony stimulating factor (G-CSF) was initiated for all patients after day 14 bone marrow biopsy, as per institutional procedure. Four of 10 patients experienced an adverse event attributed to midostaurin. Maculopapular rash was observed in 3 patients a median of 5 days after midostaurin initiation: 2 of 3 patients had a grade 2 rash and continued therapy with topical steroids; one patient had a grade 3 rash and discontinued midostaurin after 17 of 28 planned doses. A grade 1 drug fever was attributed to midostaurin in a fourth patient. Fevers persisted despite neutrophil recovery and subsequently dissipated after completion of the final midostaurin dose. Persistent FLT3 mutation was detected in 4/9 (1 not reported) day 14 bone marrow biopsies but was negative in 9/10 pts on day 28. The lone positive FLT3 result on day 28 occurred in a patient with refractory disease (>5% blasts) necessitating salvage therapy. Notably, this patient only completed 17 of 28 planned doses. All other patients exhibited a complete response (CR) on day 28. The median follow-up time was 243 days (range: 57 to 394). Nine patients are alive at the time of reporting. Six patients proceeded to allogeneic transplantation -one death was attributed to transplant-related complications, occurring in the same patient needing salvage and reduced duration midostaurin. Two patients relapsed a median of 184 days after start of induction -both FLT3-ITD positive and neither having undergone allogeneic transplant prior to relapse. Conclusions: Midostaurin in combination with idarubicin-based induction 3+7 therapy in this first case series appears to be safe. While the incidence of rash was higher (30%) than reported in RATIFY, this only resulted in discontinuation of therapy in one patient. Although patient numbers are limited, 90% achieved a FLT3 negative CR after completion of induction therapy and six patients proceeded to allogeneic transplantation. A confounding variable includes the routine use of G-CSF, which likely contributed to the shorter duration from induction to neutrophil recovery observed compared to RATIFY (23 days vs 26 days). The influence of G-CSF use on outcome is uncertain, however represents an interesting observation. A randomized, prospective trial comparing midostaurin in combination with idarubicin versus daunorubicin at both 60mg/m2 and 90mg/m2 is warranted to establish the optimal anthracycline induction therapy for FLT3 mutated AML patients. Disclosures Al-Homsi: Celyad: Membership on an entity's Board of Directors or advisory committees. Diefenbach:Denovo: Research Funding; Merck: Consultancy, Research Funding; Acerta: Research Funding; Incyte: Research Funding; Trillium: Research Funding; Millenium/Takeda: Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Genentech: Consultancy; Seattle Genetics: Consultancy, Research Funding.

We hypothesized that incremental improvements in the cyclophosphamide-doxorubicin-vincristine-prednisone (CHOP) chemotherapy regimen through optimization of drug selection, schedule, and pharmacokinetics would improve outcome in patients with large B-cell lymphomas. A prospective multi-institutional study of administration of etoposide, vincristine, and doxorubicin for 96 hours with bolus doses of cyclophosphamide and oral prednisone (EPOCH therapy) was done in 50 patients with previously untreated large B-cell lymphomas. The doses of etoposide, doxorubicin, and cyclophosphamide were adjusted 20% each cycle to achieve a nadir absolute neutrophil count below 0.5 × 109/L. The median age of the patients was 46 years (range, 20-88 years); 24% were older than 60 years; and 44% were at high-intermediate or high risk according to International Prognostic Index (IPI) criteria. There was a complete response in 92% of patients, and at the median follow-up time of 62 months, the progression-free survival (PFS) and overall survival (OS) rates were 70% and 73%, respectively. Neither IPI risk factors nor the index itself was associated with response, PFS, or OS. Doses were escalated in 58% of cycles, and toxicity levels were tolerable. Significant inverse correlations were observed between dose intensity and age for all adjusted agents, and drug clearance of doxorubicin and free etoposide was also inversely correlated with age (r = −0.54 andP2 = .08 and r = −0.45 andP2 = .034, respectively). Free-etoposide clearance increased significantly during successive cycles (P2 = .015). Lymphomas with proliferation of at least 80% had somewhat lower progression and those expressing bcl-2 had significantly higher progression (P2 = .04). Expression of bcl-2 may discriminate the recently described activated B-like from germinal-center B-like large-cell lymphomas and provide important pathobiologic and prognostic information. Dose-adjusted EPOCH may produce more cell kill than CHOP-based regimens. Dynamic dose adjustment may overcome inadequate drug concentrations, particularly in younger patients, and compensate for increased drug clearance over time.

Background: In Hodgkin lymphoma (HL) the malignant Hodgkin Reed-Sternberg (HRS) cells comprise only a small fraction of the total cellular tumor population, which orchestrate an inflammatory microenvironment of reactive cells that propagate a permissive milieu for HL growth. This HRS cell mediated immune suppression has effects that extend beyond the direct tumor microenvironment. Previously we have shown that the circulating CD4 and CD8 T cells of relapsed (R) HL patients show high expression of the receptor programmed death-1 (PD-1), and that the central memory T cells (TCM) of both newly diagnosed (ND) and R HL patients are perturbed towards chronic activation. Here we describe immune activation and perturbation in the myeloid compartment of 20 HL patients with newly diagnosed (ND) or relapsed (R) disease. Methods: Informed consent obtained from patients with ND (n= 14) or R (n=6) HL treated since January of 2013. Blood samples were drawn pre-treatment, and at sequential time-points during and after therapy. Cryopreserved PBMCs were thawed then evaluated with multi-color flow cytometry. Cells were stained with fixable viability dye (eBioscience); then stained with fluorescent-conjugated antibodies. For intracellular staining, cells were fixed and permeabilized using FOXP3 staining kit (eBioscience) then stained with intracellular antibodies. Stained cells were analyzed using LSRII flow cytometer (BD Bioscience) and FlowJo software (Tree Star). The following anti-human antibodies were used to analyze myeloid subsets: CD3, CD19, CD16, HLA-DR, CD14, CD303, CD141. Monocytes are defined as CD14+HLA-DR+CD3-CD19-, plasmacytoid dendritic cells (pDCs) as: CD303+HLA-DR+CD14-CD3-CD19-CD16-, and dendritic cells (DCs) as: HLA-DR+CD141+CD14-CD303-CD3-CD19-CD16-(Biolegend). Singlet cells were gated based on forward and side scatter properties. Dead cells were excluded based on viability dye. Patient samples were compared to normal controls matched for age and sex (n=20). Results: The median HL patient age was 38 (range: 20-90 years). Twelveof the HL subjects were male and 8 were female. All but 1 of the ND HL patients were treated with upfront ABVD +/- consolidative radiation. Two out of 6 R patients had prior allogeneic stem cell transplant (alloSCT); they were not on immunosuppression. Thirteen patients (12 ND, 1 R) responded to therapy (11 CR and 2 PR); 4 patients (1 ND, 3 R) progressed on therapy or had stable disease, 2 patients do not have disease status confirmed. We observed a decrease in the proportion of systemic circulating plasmacytoid dendritic cells (pDCs), but not monocytes or DCs, in HL patients at baseline compared to healthy controls (Figure 1). Interestingly, the ratio of both DCs and pDCs to monocytes were greatly disturbed in HL patients compared to healthy subjects (Figure 2). After a single cycle of chemotherapy we noted an increase in pDCs, and a decrease in the ratio of monocytes to both DCs and pDCs and of DCs to pDCs in treated patients. Together these findings suggest a systemic imbalance within the monocyte-DC-pDC axis in HL patients compared to normal healthy controls, which appears to normalize after one cycle of treatment with chemotherapy. Conclusion: HL patients have evidence of perturbation in the systemic myeloid compartment, suggesting that the impact of HRS cells on their microenvironment may have systemic as well as local effects that extend to the myeloid compartment and to antigen presentation, as well as to T cell phenotypes. This underscores the rationale for understanding the role of systemic immune dysfunction in HL, and for the use of immune targeted therapies in HL. Correlation with clinical data, and functional studies to further characterize this immune dysregulation are ongoing. Disclosures Diefenbach: Merck: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Gillead: Equity Ownership; Seattle Genetics: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Research Funding. Martin:Gilead: Consultancy, Other: travel, accommodations, expenses; Janssen: Consultancy, Honoraria, Other: travel, accommodations, expenses; Celgene: Consultancy, Honoraria; Novartis: Consultancy; Teva: Research Funding; Acerta: Consultancy. Ruan:Seattle Genetics: Consultancy, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; Pharmacyclics, LLC, an AbbVie Company: Research Funding, Speakers Bureau; Janssen: Research Funding.

Anti-B-blocked ricin (anti-B4-bR) combines the specificity of the anti- B4 (CD19) monoclonal antibody with the protein toxin “blocked ricin.” In blocked ricin, affinity ligands are attached to the ricin B-chain to attenuate its lectin binding capacity. In a phase I trial, Anti-B4-bR was administered by 7-day continuous infusion to 12 patients in complete remission after autologous bone marrow transplantation (ABMT) for relapsed B-cell non-Hodgkin's lymphoma (NHL). Patients were treated at 20, 40, and 50 micrograms/kg/d for 7 days. Potentially therapeutic serum levels could be sustained for 3 to 4 days. The maximum tolerated dose was 40 micrograms/kg/d for 7 days (total 280 micrograms/kg). The dose-limiting toxicities were reversible grade IV thrombocytopenia and elevation of hepatic transaminases. Mild capillary leak syndrome was manifested by hypoalbuminemia, peripheral edema (4 patients), and dyspnea (1 patient). Anti-immunotoxin antibodies developed in 7 patients. Eleven patients remain in complete remission between 13 and 26 months post-ABMT (median 17 months). These results show that Anti-B4- bR can be administered with tolerable, reversible toxicities to patients with B-cell NHL in complete remission following ABMT.

Anti-B4-blocked Ricin (Anti-B4-bR) is an immunotoxin comprised of the anti-B4 monoclonal antibody (MoAb) and the protein toxin “blocked ricin.” The anti-B4 MoAb is directed against the B-lineage-restricted CD19 antigen expressed on more than 95% of normal and neoplastic B cells. Blocked ricin is an altered ricin derivative that has its nonspecific binding eliminated by chemically blocking the galactose binding domains of the B chain. In vitro cytotoxicity studies demonstrate that the IC37 of Anti-B4-bR is 2 x 10(-11) mol/L compared with 4 x 10(-12) mol/L for native ricin. A phase I dose escalation clinical trial was conducted in 25 patients with refractory B-cell malignancies. Anti-B4-bR was administered by daily 1-hour bolus infusion for 5 consecutive days at doses ranging from 1 microgram/kg/d to 60 micrograms/kg/d. Serum levels above 1 nmol/L were achieved transiently in the majority of patients treated at the maximum tolerated dose of 50 micrograms/kg/d for 5 days for a total dose of 250 micrograms/kg. The dose-limiting toxicity was defined by transient, reversible grade 3 elevations in hepatic transaminases, without impaired hepatic synthetic function. Minor toxicities included transient hypoalbuminemia, thrombocytopenia, and fevers. Human antimouse antibody and human anti-ricin antibody were detected in nine patients. One complete response, two partial responses, and eight mixed or transient responses were observed. These results show the in vitro and in vivo cytotoxicity of Anti-B4-bR and indicate that this immunotoxin can be administered as a daily bolus infusion for 5 days with tolerable, reversible toxicity.

Background Among therapies for immune thrombocytopenia (ITP) that achieve long unmaintained remissions, distant relapse rates have not been compared. Specific Aims To compare relapse rates following umaintained remissions exceeding 1 year in patients with ITP treated with steroids versus anti-Rh(D) with/without steroids versus rituximab with/without steroids. Methods Institutional Review Board approval was obtained. The charts of 52 consecutive patients with ITP followed in a hematology practice between 07/01/03 and 06/30/13 were reviewed. Remission was defined as a platelet count > 100,000/µL. Patients with the aforementioned treatment and remission characteristics were identified. Steroids were administered alone as dexamethasone 40 mg PO daily for 4 days. Anti-Rh(D) was administered as 75 mcg/kg IV with or without prednisone 60-85 mg PO daily tapered over 4 weeks or dexamethasone 40 mg PO daily for 4 days. Rituximab was administered as 375 mg/m2 IV weekly for 4 weeks or as a single dose with or without dexamethasone 40 mg PO daily for 4 days. Relapse was defined as a platelet count < 30,000/µL. Statistical calculations included ANOVA to compare demographics and chi-square contingency table analysis to compare distant relapse rates (http://www.physics.csbsju.edu/stats). Results There were 16 unmaintained remissions exceeding 1 year in 13 patients following treatment with steroids, anti-Rh(D) with/without steroids, or rituximab with/without steroids. Mean age on presentation, sex ratio, duration of ITP prior to therapy, and duration of follow-up after therapy were similar for the 3 groups (Table 1). Distant relapse rates were 100%, 14.2%, and 66.7% for the steroid, anti-Rh(D) with/without steroid, and rituximab with/without steroid groups, respectively, P = 0.03 (Table 2). Conclusion Among patients who achieve long unmaintained remissions of ITP with steroids, anti-Rh(D) with/without steroids, or rituximab with/without steroids, those treated with anti-Rh(D)-based therapy are the most likely to have extended remission. Disclosures: Off Label Use: Rituximab therapy of ITP.

Background Anti-Rh(D) (Cooper Blood 2002), rituximab (Godeau Blood 2008), and dapsone (Sharma ASH 2006) have been shown to be splenectomy-sparing agents for the treatment of immune thrombocytopenia (ITP). In these studies, patients treated with anti-Rh(D) and rituximab were followed for 2 years; patients were treated with dapsone for 6 to 48 months. Specific Aims To determine long-term outcomes in unsplenectomized patients with ITP treated with anti-Rh(D), rituximab, or dapsone. Methods Institutional Review Board approval was obtained. The charts of 52 consecutive patients with ITP followed in a hematology practice between 07/01/03 and 06/30/13 were reviewed. Response was defined as a platelet count (PLT) > 100,000/µL. Relapse was defined as PLT < 30,000/µL. Results Twenty-one patients have been treated with anti-Rh(D), rituximab, or dapsone. Some patients treated with anti-Rh(D) or rituximab received brief concurrent steroids. For the entire cohort the mean duration of ITP was 3.50 years (y), range 0.008 to 19 y and the mean duration of follow-up was 3.84 y, range 0.10 to 8.81 y. Among 16 patients who received anti-Rh(D), 85% responded to the initial infusion. Among the 10 responders who have had follow-up exceeding 1 year, 4 have had a sustained response following the first infusion (mean follow-up duration 3.35 y, range 2.66 y to 5.98 y; six have relapsed, all have responded to retreatment. Among 7 patients who received rituximab, 71% responded to the initial treatment. All 5 responders have had follow-up exceeding 1 year, among whom 2 have had a sustained response following the first infusion (mean follow-up duration 2.24 y, range 2.19 y to 6.67 y); three have relapsed, all have responded to retreatment. Among 3 patients who received dapsone, 66.7% responded; the response is partial in 1 patient with PLT 50,000/µL (mean follow-up duration 3.94 y, range 0.11 y to 8 y). Toxicities have included: with anti-Rh(D) 1 case each of grade II uterine pain, grade II chills, and grade II non-cardiac chest pain; with rituximab 1 case of grade II back pain. Therapeutic switches have included 2 cases of anti-Rh(D) to rituximab and 1 case each of anti-Rh(D) to rituximab to dapsone, anti-Rh(D) to rituximab to dexamethasone, anti-Rh(D) to romiplostim, and dapsone to rituximab. No patients have undergone splenectomy. Conclusion Anti-Rh(D), rituximab, and dapsone are splenectomy-sparing therapies for ITP over an extended period. For patients treated with anti-Rh(D) or rituximab who relapse, retreatment is effective. Disclosures: Off Label Use: Rituximab and dapsone therapies of ITP.

Abstract 3343 Background Anti-Rh(D) is an immune globulin against the Rh group and is used for the treatment of immune thrombocytopenia (ITP). A 75 μcg/kg dose induces an overnight response in 92% of patients and a median response duration of 46 days. More than 50% of patients treated intermittently with anti-Rh(D) continue to require therapy after one year. Danazol, an attenuated androgen, has a longer onset of action (median 3.1 months), but has steroid-sparing activity in ITP. We conducted a phase II study to determine whether danazol could lower the requirement for recurrent use of anti-Rh(D). Methods Twenty-six patients with ITP were enrolled. Treatment consisted of daily danazol 600 mg PO in combination with anti-Rh(D) 75 μg/kg IV on day 1, repeated whenever the platelet count (PLT) fell below 30,000/μL. After 1 year, danazol was reduced to 400 mg daily for 3 months then to 200 mg daily for 9 months. Chi-square analysis was used to compare requirement for just a single infusion of anti-Rh(D) with that in historical controls treated with anti-Rh(D) alone. The student t-test was used to compare rate of anti-Rh(D) usage over time in both groups. Results were analyzed by intent-to-treat. Results Patients' median age was 45 years (range 23–83 years). The male:female ratio was 1.9:1. Eleven (42%) were human immunodeficiency virus (HIV)-positive. Median time from diagnosis of ITP to enrollment was 10.5 months. Median PLT at enrollment was 15,000/μL. Nine (35%) of patients had received a median of one prior infusion of anti-Rh(D). Median duration of follow-up was 28.12 months. Nine (34.6%) of 26 study patients required just a single infusion of anti-Rh(D) in comparison with 1 (3.6%) of 28 control patients, p = 0.003. There was no correlation of age, sex, HIV status, and number of months from diagnosis of ITP with need for a single infusion of anti-Rh(D). At 1 year of follow-up, there was a 20% reduction in anti-Rh(D) use in comparison with historical controls: mean number of infusions per month 0.36 for study patients vs. 0.43 for controls, t = 0.9066, p > 0.05. Platelet responses in HIV-positive patients were not attributable to antiviral treatment. Study therapy was well-tolerated. Two patients stopped danazol due to intolerable adverse effects (grade III acne, grade II myalgias, grade I alopecia.) Conclusion Danazol demonstrates anti-Rh(D)-sparing activity. Greater than 1/3 of patients treated with the combination require just a single infusion of anti-Rh(D). The cost of an anti-Rh(D) infusion ($ 1500.00) is nearly one-half the cost of daily danazol for 1 year ($ 3600.00). Historical controls treated with anti-Rh(D) alone required a median of 5 infusions per year whereas patients treated with the combination in this study required a median of 2 infusions per year. This reduction in anti-Rh(D) use would have been associated with cost savings. This study was underpowered to detect a significant difference in rate reduction of anti-Rh(D) use over time. Future studies should evaluate the use of this combination in a larger population. Disclosures: No relevant conflicts of interest to declare.

Background: The goal of therapy of immune thrombocytopenic purpura (ITP) is to produce and maintain a platelet (PLT) count greater than 30,000/μL. While anti-Rh(D) produces a rapid PLT rise (43,000/μL median overnight increase), the median duration of response is short (46 days). More than 50% of patients treated intermittently with anti-Rh(D) continue to require therapy after one year. In contrast to anti-Rh(D), the time to response and duration of response with danazol are long (median 3.1 months, 119 months, respectively). Both agents are alternatives to splenectomy. We hypothesize that danazol is an anti-Rh(D)-sparing agent and that the addition of danazol to anti-Rh(D) may decrease the requirement for anti-Rh(D) and obviate the need for continued anti-Rh(D) after one year. Methods: This is a single arm, non-randomized, phase II trial, utilizing a single stage design to compare rates of discontinuance of anti-Rh(D) at the end of one and two years. A total of 26 patients will be recruited. Treatment consists of daily danazol 600 mg in combination with anti-Rh(D) 75 μg/kg IV on day 1, to be repeated whenever PLT falls below 30,000/μL. After 1 year, danazol will be reduced to 400 mg daily for 3 months then to 200 mg daily for 9 months. Results: Seven patients have been enrolled to date (5 men/2 women). The median patient age is 43 years (range 22–51 years). The clinical course on study is summarized in Table 1. Five patients are evaluable for efficacy. Four patients have required a median of 1.5 anti-Rh(D) infusions during a median study enrollment of 43.5 weeks. Three patients have had sustained responses following the initial infusion of anti-Rh(D) noted at 14 - 35 weeks of follow-up; a second course of anti-Rh(D) was required in one of these patients at 40 weeks of follow-up. One patient has developed progressive disease. Two patients were removed from protocol therapy due to danazol toxicities; they have maintained PLT greater than 30,000/μL 6 - 35 weeks after discontinuation. No patients were noncompliant with therapy. No treatment-related deaths occurred. Conclusion: The combination of anti-Rh(D) and danazol is a novel regimen for patients with ITP. While anti-Rh(D) provides an immediate effect, danazol provides maintenance therapy. The duration of response after the initial anti-Rh(D) infusion in three patients in this study appears longer than that in patients treated with anti-Rh(D) alone, but further accrual and follow-up will be required to define toxicity and efficacy. Efficacy of anti-Rh(D) and danazol as of 07/27/06 Pt. Pt. Prior therapy Duration of danazol therapy (wks) # of anti-Rh(D) infusions administered Toxicities Comments 1 Prednisone, Anti-Rh(D)x1 47 2 Second course of anti-Rh(D) was required 40 weeks after the initial course. 2 Dexamethasone, IVIG, Anti-Rh(D)x3 14 1 Gr 1 alopecia, Gr 2 myalgias Disenrolled due to toxicity. Has maintained PLT > 30,000/ μL 6 weeks after disenrollment. 3 None 43 1 4 Dexamethasone 44 9 5 Dexamethasone, IVIG 6 4 Gr 2 anemia Disenrolled due to progressive disease (retroperitoneal bleed). 6 Prednisone 6 1 Disenrolled due to technical limitations (lack of IV access). 7 Dexamethasone 3 1 Gr 3 acne Disenrolled due to toxicity. Has maintained PLT > 30,000/ μL 35 weeks after disenrollment.

Diffuse large B-cell lymphoma (DLBCL) is subdivided by microarray into germinal center B-cell (GCB) and activated B-cell (ABC) subtypes. These subtypes have distinct pathways of lymphomagenesis and outcome with 5-year survival of 59% and 31%, respectively, using CHOP. A molecular predictor showed high tumor proliferation and the ABC subtype are associated with a poor outcome. We showed DA-EPOCH overcomes the adverse effects of high tumor proliferation (BLOOD 99:2685, 2002) and rituximab overcomes the adverse effects of BCL-2, which is associated with the ABC subtype (Proc Am Soc Heme 102:abstr 356, 2003). To assess DA-EPOCH-R with G-CSF in GCB and ABC subtypes, we employed a validated immunohistochemistry (IHC) algorithm (BLOOD: 103:275, 2004). GCB was defined as CD10+ or BCL-6+ and MUM1-. ABC was defined as CD10- and BCL-6- or CD10-, BCL-6+ and MUM1+. Eligibility includes de novo DLBCL, no prior chemotherapy, any PS, HIV- and stage I thymic DLBCL > 5 cm and all stage II-IV. Characteristics of 77 enrolled pts include median (range) age 48 (12-85) and PS 1 (0-3); stage III/IV 48 (62%); and LDH > nl 42 (55%); extranodal sites >1 27 (35%); and HI/H IPI 26 (34%). Including all 77 pts, response is CR/CRu 72 (94%). At the median (range) follow-up of 28 (4-60) mos, PFS is 82% and OS is 83% (Fig 1). DFS is 87% at 28 mos with no relapses beyond 18 mos. PFS at 28 mos according to IPI risk is 95% for 0-2 factors and 54% for 3-5 factors. Using IHC, 44 pts were subdivided in GCB (35) and ABC (9) subtypes. The proportion of the ABC subtype vs. GCS is low and likely due to incomplete IHC for the ABC subtype. Specifically, 16 cases that were CD10- did not have adequate BCL-6 and/or MUM1 IHC for categorization. PFS at 28 mos for the GCB and ABC subgroups were 84% and 62%, respectively (Fig 2). DA-EPOCH-R employs a pharmacodynamic design where doses of doxorubicin, etoposide and cyclophosphamide are adjusted (i.e. normalized) to achieve neutrophil nadirs < 500/mm3. PK analysis shows that clearance of doxorubicin and etoposide varies by up to 1 log and that dose-adjustment compensates for these differences. This approach increases dose intensity in pts with high drug clearance while limiting toxicity. Based on 451 cycles, the target nadir ANC < 500/mm3 was achieved on 277 (61%) cycles but with 67 (15%) episodes of fever/neutropenia. Non-hematological toxicity was low and there were 2 treatment related deaths. These results suggest DA-EPOCH-R is highly effective in all subtypes of de novo DLBCL. A phase III study comparing R-CHOP and DA-EPOCH-R with microarray analysis of tumor biopsies is under development. Figure Figure Figure Figure