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Background: Recurrent/refractory (r/r) DLBCL presents a major treatment challenge, especially in the setting of patients who are ineligible for, or relapsing after, potentially curative treatments such as autologous stem cell transplant (ASCT) and chimeric antigen receptor T cell (CAR-T) therapy. Efficacious treatment options that are well-tolerated and easily accessible in this population represent a critical unmet medical need. DPX-Survivac is a targeted T cell activation therapy against cancers expressing survivin. Survivin plays an essential role in cancer biology and represents a target of choice to disrupt tumour progression. DPX-Survivac's mechanism of action relies on its ability to generate robust and durable survivin-specific T cells that migrate to, infiltrate and kill tumour cells. DPX-Survivac is administered with intermittent, low dose cyclophosphamide (CPA) used as an immunomodulator. In nonclinical studies, treatment with DPX-Survivac increases PD-L1 and PD-1 expression providing the rationale for combination with pembrolizumab. Methods: "SPiReL" is a Phase 2 non-randomized, open label, efficacy and safety study. Subjects with r/r DLBCL with confirmed survivin expression are eligible for participation. Subjects must also be ineligible for potentially curative therapy. The treatment and testing regimen is shown in the figre below. The primary objective of SPiReL is to document a minimum Objective Response Rate (ORR) of 24% (in 6/25 subjects) using the modified Cheson criteria (2007). Secondary objectives include safety, duration of response and time to next treatment. Exploratory endpoints include T cell response, tumour immune cell infiltration, and biomarker analysis. Results: At data cut-off, 22 subjects have been enrolled in the study. The median age is 75.5 years (50-82). Thirteen of 22 subjects (59.1%) are GCB sub-type, 8 subjects (36.4%) are non-GCB and 1 subject (4.5%) has primary cutaneous DLBCL, leg type. The median number of prior therapies is 2 (1-7), with 4 subjects having previously undergone ASCT and 5 subjects with transformed disease. Of the 22 enrolled subjects, 8 are not evaluable per protocol for clinical efficacy due to early disease progression. Four subjects are active on treatment, 3 of which have not yet reached the first time point for assessment. Clinical outcome was analyzed for the Full Analysis Set (FAS) (n=19) and in the Per Protocol (PP) analysis (n=11). Of 11 subjects in the PP, 7 subjects (63.6%) have achieved an objective response, meeting the study's primary endpoint; 3 subjects (27.3%) with a CR and 4 subjects (36.4%) with a PR. Three subjects (27.3%) achieved SD and thus clinical benefit was demonstrated in 10/11 (90.9%) of evaluable subjects. Two subjects (18.2%) have completed the 1 year treatment period, 8 subjects (72.7%) discontinued treatment due to disease progression, and 1 subject (9.2%) discontinued treatment due to an unrelated Adverse Event (AE). Including the entire FAS, the objective response rate was 7/19 (36.8%). This treatment combination is well-tolerated, only 11% of Treatment Related AEs (TRAEs) were assessed as Grade 3 or higher. The majority of the Grade 1 and 2 TRAEs were Injection Site Reactions (ISRs) related to DPX-Survivac. Ten serious AEs were reported, of which 3 were considered related to study treatment. No subjects have discontinued study treatment due to a TRAE. Analyses of peripheral blood T cell responses to survivin by ELISpot assay shows that 7/7 subjects who achieved an objective response have survivin-specific T cell response. One subject with PD also showed a survivin-specific T cell response, supporting the mechanism of action and the role of DPX-Survivac in anti-tumor activity. Summary: DPX-Survivac and low dose CPA in combination with pembrolizumab, demonstrates promising clinical activity in recurrent/refractory DLBCL with 10/11 (90.9%) of evaluable subjects deriving clinical benefit with minimal toxicity. The primary endpoint of this study has been reached with 7/11 (63.6%) of evaluable subjects achieving an objective response warranting further exploration of DPX-Survivac in this population. Enrollment is continuing to further define the patient population most likely to benefit from this well-tolerated therapy. Figure 1 Disclosures Bence-Bruckler: Merck: Membership on an entity's Board of Directors or advisory committees. Forward:Seattle Genetics: Research Funding; IMV: Research Funding; Merck: Research Funding; Astellas: Research Funding; Servier: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; IMV: Membership on an entity's Board of Directors or advisory committees; Calgene: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AstraZeneca: Membership on an entity's Board of Directors or advisory committees. Stewart:Roche: Honoraria; Janssen: Honoraria; Abbvie: Honoraria; Gilead: Honoraria; Celgene: Honoraria; Amgen: Honoraria; Sandoz: Honoraria; Novartis: Honoraria; AstraZeneca: Honoraria; Teva: Honoraria. Bramhecha:IMV Inc.: Current Employment. Conlon:IMV Inc.: Current Employment. OffLabel Disclosure: Keytruda (pembrolizumab). Indicated for use in melanoma, nonÃÆ'Ã'¢Ã¢ââ'¬Å¡Ã¢â€šâ'¬ÃƒÂ¢Ã¢â€šâ'¬Ã…“small cell lung cancer (NSCLC), head and neck squamous cell cancer (HNSCC), classical Hodgkin lymphoma (cHL), primary mediastinal B-cell lymphoma (PMBCL), urothelial carcinoma, microsatellite instability-high (MSI-H) or a mismatch repair deficient (dMMR) solid tumor, gastric or gastroesophageal junction (GEJ) adenocarcinoma, squamous cell carcinoma of the esophagus, cervical cancer, hepatocellular carcinoma, Merkel cell carcinoma (MCC), and renal cell carcinoma (RCC).

Introduction: Recurrent/refractory diffuse large B cell lymphoma (DLBCL) remains an area of unmet clinical need. Although autologous stem cell transplant (ASCT), and more recently chimeric antigen receptor (CAR) T cell therapy, have made a significant impact, many patients are ineligible for these treatments due to advanced age, comorbidities, or financial or geographical limitations, or they have relapsed following these treatments. Survivin, a novel target in DLBCL, is overexpressed in approximately 60% of untreated DLBCL with higher rates in relapsed disease. DPX-Survivac, a T cell activating therapy, elicits a strong and prolonged immune response against tumors expressing survivin. DPX-Survivac is combined with pembrolizumab, an anti-PD-1 checkpoint inhibitor, and intermittent low dose cyclophosphamide (CPA), used for an immunomodulatory effect. Pre-clinical studies have demonstrated increased efficacy of this DPX-based drug combination in controlling of tumor growth in murine tumor models, resulting in improved survival (Weir et al. J Immunother Cancer 2016). Methods: "SPiReL" is a Phase 2 non-randomized, open label, efficacy and safety study. Subjects with recurrent/refractory DLBCL with confirmed survivin expression are eligible for participation. Participants must also be ineligible for curative therapy. Study treatment includes administering two doses of 0.5 mL of DPX-Survivac 3 weeks apart followed by up to six 0.1 mL doses every 8 weeks. Intermittent low dose cyclophosphamide is administered orally at 50 mg twice daily for 7 days followed by 7 days off. Pembrolizumab 200 mg is administered every 3 weeks. Study participants continue active therapy for up to one year or until disease progression, whichever occurs first. The primary objective is to document the response rate to this treatment combination using modified Cheson criteria. Secondary objectives include duration of response and safety. Exploratory endpoints include T cell response, tumor immune cell infiltration, and gene expression analysis. Enrollment is ongoing with a goal of up to 25 subjects in this national, multi-center study. Trial design is shown below the text. Results: At the time of data cut-off, 23 subjects have been screened and 12 have been enrolled. The demographics of enrolled subjects include: median age is 75.5 years (50-82), with 3 male subjects participating. The median number of prior therapies is 2.5 (1-6), with 4 subjects having previously undergone ASCT. The median time from diagnosis to screening is 31 months (8-151). Of the 12 enrolled subjects, 3 were not evaluable for clinical efficacy due to early disease progression. One subject has not yet reached the first time point for assessment. In the Per Protocol analysis of 8 evaluable subjects ("PP", N = 8); as compared to a Full Analysis Set ("FAS", N = 11); 7 of 8 subjects (PP = 87.5%) demonstrated clinical benefit, including 6 with tumor regressions (FAS = 63.6%); 2 subjects (PP = 25%) achieved a complete response, 1 of whom has completed the 1 year study period (FAS = 18.2%); 3 subjects (PP = 37.5%) had a partial response (FAS = 27.3%); and 3 (PP = 37.5%) had stable disease remaining on study for 4, 6 and 8 months (FAS = 27.3%). This treatment combination is well-tolerated with mostly grade 1 and 2 adverse events reported; a single grade 3 maculopapular rash and 1 case of grade 4 cytopenia has been observed. Of the 23 screened subjects, survivin expression analysis was performed on 18 subjects. All of the 18 samples analyzed were survivin positive, with a range of 60%-100% of lymphoma cells expressing survivin. Preliminary analysis of peripheral blood T cell responses to survivin by ELISPOT assay shows 2 of 4 subjects had strong survivin-specific T cell responses. Both of these subjects had clinical responses. Summary: DPX-Survivac, in combination with pembrolizumab and low dose cyclophosphamide, demonstrates clinical activity and a favorable safety profile in recurrent/refractory DLBCL with 87.5% of evaluable subjects deriving clinical benefit, including 2 complete responses (25%) and 3 partial responses (37.5%) to date. Enrollment is continuing to further explore the synergistic effect of this combination in this population. Figure Disclosures Berinstein: Merck: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Stewart:Merck: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. OffLabel Disclosure: Keytruda (pemroblizumab) is approved for use to treat Melanoma, Metastatic, Non-Small Cell Lung Cancer, Head and Neck Cancer, Hodgkin's Lymphoma, Urothelial Carcinoma, Gastric Cancer, Cervical Cancer, Hepatocellular Carcinoma, Merkel Cell Carcinoma, Renal Cell Carcinoma, and Small Cell Lung Cancer. In this trial, it is being used to treat Diffuse Large B-Cell Lymphoma.

Background: Although the natural history of follicular lymphoma is indolent with a median overall survival of about 12-15 years, the disease is heterogeneous. The 5 and 10 year overall survival (OS) of low, intermediate and high risk FLIPI is 91%, 78% and 53% and 71%, 51% and 36% using standard rituximab-based treatment. 5-year progression-free survival (PFS) is 80%, 70% and 48% respectively. Methods: Based upon this we conducted an investigator-initiated single-centre Phase II trial of intensified therapy with CHOP-R followed by (90)Y ibritumomab tiuxetan consolidation and 24 months of rituximab maintenance as treatment for patients with intermediate and high-risk newly diagnosed symptomatic follicular lymphoma. 33 patients were enrolled. Results: The addition of (90)Y ibritumomab tiuxetan was well tolerated but resulted in asymptomatic grade 3 or 4 thrombocytopenia and neutropenia in11-36% and 10-24% of patients between weeks 2-8 post (90) Y. After 9 years of follow-up (median follow-up 61 months) the 0S for intermediate and high risk FLIPI was 95% and 78%. The 5 year PFS was 79% and 64% for intermediate and high risk FLIPI, respectively. Responses at three months post consolidation were as follows: 3/33 (9%) achieved CR, 25/33(76%), achieved CRU, 1/33(3%) had PR, and 1/33(3%) had PD. Three patients did not receive (90)Y ibritumomab tiuxetan due to disease progression 2/33(6%), or death 1/33(3%). Of 19 patients who had a molecular marker for their lymphoma, 18 (95%) achieved molecular remissions in peripheral blood with CHOP-R therapy. Nine (47%) of these patients have been recently assessed for MRD and remain in molecular remission. The therapy resulted in decreased levels of IgG, IgM and IgA below the lower normal level in 33%, 40% and 23% of patients respectively post therapy. These levels did not recover in most of these patients. B cells were depleted to undetectable levels during therapy including rituximab maintenance. In 18 evaluable patients only 11 recovered normal B cell counts post maintenance rituximab. There was no correlation between normal B-cell recovery and Ig levels. Many patients with low or no B cell counts had normal IgG levels, whereas some patients who regained normal B cell counts were still unable to reach normal Ig levels. No patient developed human anti-mouse antibody. Immunity to measles, mumps, or rubella was retained post therapy. Patients did not have significant infections or opportunistic infections (although 2 developed Grade 1 shingles post (90)Y ibritumomab tiuxetan) and none required IVIG. Conclusions: We conclude that this intensified regimen is highly active in cyto-reducing lymphoma in high and intermediate risk FLIPI follicular lymphoma patients. The toxicity is tolerable although a significant percentage of patients will end up with persistent asymptomatic reductions in B cells and serum Ig. Only randomized trials will determine whether this regimen enhances outcome over standard of care in this higher risk follicular lymphoma population. References: 1.Examination of the follicular lymphoma international prognostic index (FLIPI) in the National LymphoCare study (NLCS): a prospective US patient cohort treated predominantly in community practices. Nooka AK, Nabhan C, Zhou X, Taylor MD, Byrtek M, Miller TP, Friedberg JW, Zelenetz AD, Link BK, Cerhan JR, Dillon H, Sinha R, Shenoy PJ, Levy D, Dawson K, Hirata JH, Flowers CR. Ann Oncol. 2013 Feb;24(2):441-8. doi: 10.1093/annonc/mds429. Epub 2012 Oct 5 2.Validation, revision and extension of the Follicular Lymphoma International Prognostic Index (FLIPI) in a population-based setting. van de Schans SA, Steyerberg EW, Nijziel MR, Creemers GJ, Janssen-Heijnen ML, van Spronsen DJ. Ann Oncol. 2009 Oct;20(10):1697-702. doi: 10.1093/annonc/mdp053. Epub 2009 Jun 23. PMID: 19549712 Disclosures Buckstein: Novartis: Honoraria; Celgene: Honoraria, Research Funding.

Diffuse large cell lymphomas are aggressive tumors of B-cell origin. In some cases they arise from low-grade follicular lymphomas carrying the t(14;18) translocation, an event that leads to the overexpression of the BCL-2 gene product. More frequently, however, they lack the t(14;18) translocation. Rearrangements of the c-MYC proto-oncogene and mutations of the p53 tumor suppressor gene have also been documented in these lymphomas. This study examines the extent to which alterations in the BCL-2, c-MYC, and p53 genes co-exist within individual lymphomas. Eight diffuse large cell lymphoma cell lines and 11 diffuse large cell lymphoma tumors were assessed for genetic alterations in these three genes. Our results indicate that there is a heterogeneity in the oncogene/suppressor gene profile among diffuse large cell lymphomas. Two cell lines and one tumor carried alterations in all three genes, one cell line carried alterations of c-MYC and p53, and one primary tumor and one cell line carried p53 mutations and the t(14;18). Single alterations of BCL-2 and p53 were also observed. Another cell line had no alterations in any of these genes. The heterogeneity indicates that varied mechanisms may be involved in the generation of diffuse large cell lymphomas.

Several genetic mechanisms have been shown to diversity the expressed antibody repertoire of committed B lymphocytes. These include somatic hypermutation, V gene replacement, and ongoing gene rearrangement. These mechanisms may be operational at discrete points in the B-cell differentiation pathway and may generate idiotypic diversity in various malignant B-cell tumors. Hypermutation of the Ig variable region has been shown to occur in follicular lymphoma, but not in pre-B cell acute lymphoblastic leukemia, Burkitt's lymphoma, chronic lymphocytic leukemia, or myeloma. To study hypermutation in a large cell lymphoma, we use a polymerase chain reaction-based approach, employing consensus VH and JH primers, to clone and sequence rearranged Ig heavy chain variable regions. Neither tumor cells immortalized in rescue fusions nor idiotypic variants of a tumor-derived cell line generated through ongoing lambda light chain gene rearrangements show any significant number of variable region mutations. Thus, at the in vivo stage of B- cell differentiation from which this large cell lymphoma arose, Ig variable region hypermutation was not occurring, nor did it occur during propagation in vitro of these tumor cells. Thus, the window of hypermutation in malignant B-cell tumors is more precisely defined, which may have clinical implications for diagnostic and therapeutic approaches directed at the Ig variable region.

Background Dose-modified(dm) CODOX-M/IVAC is commonly used for Burkitt lymphoma (BL) and overlap aggressive B cell lymphomas (B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma (DLBCL) and BL, previously Burkitt-like lymphoma-BLL), with high cure rates, but significant toxicity. The concurrent use of rituximab with CODOX-M/IVAC is relatively new in both BL and BLL. Methods: We undertook this retrospective study of BL and BLL patients treated at a large quaternary care cancer center with dmCODOX-M/IVAC +/-rituximab(R) in order to a) determine ‘real-world' toxicity and survival rates, b) examine the effects of BL versus intermediate histology diagnoses, and c) evaluate the effects of rituximab (R) use. Patients diagnosed between January 2006-December 2012 as having either Burkitt lymphoma or atypical Burkitt/Burkitt-like lymphoma (WHO Classification 2001), or B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma and BL (WHO 2008) and who had received the dmCODOX-M/IVAC regimen,+/- R within 2 months of their diagnosis, were identified. In May 2008, we adopted the practice of adding R 375 mg/m2 to dmCODOX-M/IVAC in non-HIV patients due to emerging data from other centers. In 2012, our approach to BLL was altered, by adding 1-4 cycles of R-CHOP after the dmCODOX-M/IVAC protocol completion, to maximize anthracycline and rituximab delivery. The primary outcomes of interest were 2-year overall (OS) and progression-free survival (PFS). Univariate and multivariable Cox proportional hazard models of OS and PFS were constructed. Results: 31 patients with BL (n=21) or BLL (n=10) were included. The median age was 45, and 90% were high risk. 94% of patients displayed a MYC translocation (t(8;14)) by FISH cytogenetics. Additionally, 8 (26%) patients were BCL2+ by immunohistochemistry (IHC). Three were also positive for the t(14;18) (double-hit lymphomas), and one was unknown. Because of the diagnostic uncertainty (pending MYC status), 10 (32%) began treatment with chemotherapy other than dmCODOX-M/IVAC (mostly 1-2 cycles of CHOP+/-R). The median time from diagnosis to first chemotherapy was 13 days (1-58) and from first chemo to first HDMTX 10.5 days (0-57). In total, 22 (71%) patients received at least one infusion of rituximab at some point during first line treatment with a median of 3 infusions (range 1-7). Five of 10 BLL patients (50%) received a median of 2 additional R-CHOP cycles (range 1-4). In the patients who received R-CHOP first vs. those who did not, the median time from diagnosis to the first chemotherapy was 9 days (range: 1-21) compared with 17 days (range: 0-58) (p=0.29). Despite GCSF use, 24 patients (77%) experienced at least one episode of febrile neutropenia post dmCODOX-M and 12 patients (39%) post dmIVAC. The 2-year overall survival (OS) and progression-free survival (PFS) were 69% and 62%, respectively, with no differences between BL and BLL (Figure 1). When BLL was separated by whether R-CHOP was given, there were still no differences in OS between BL, BLL (“no R-CHOP after”) and BLL (“R-CHOP after”) (2-yr OS 64% vs. 60% vs. 100%, p=0.38; Figure 1). Patients who received rituximab had superior OS (2-yr OS 86% vs. 33%, p=0.0013; Figure 1) compared with those that did not. Starting with CHOP-R versus dmCODOX-M was associated with an OS benefit (2-yr OS 100% vs. 53%, p=0.009; Figure 1), despite no difference in OS whether time from diagnosis to first chemotherapy was ≥ or < 10 days (p=0.96) By multivariable analysis, rituximab use was significantly associated with improved OS (HR 0.15; 95%CI 0.04-0.56) and PFS (HR 0.05; 95%CI 0.01-0.32). Conclusion The addition of rituximab to dmCODOX-M/IVAC improves overall and progression free survival in both BL and BLL and is recommended. BLL is a challenging histology to treat with inferior outcomes to DLBCL when treated similarly. The use of an aggressive regimen such as R-dmCODOX-M/IVAC in BLL can yield promising results that are comparable to BL. Disclosures: No relevant conflicts of interest to declare.

Background NCIC CTG LY.12 was an international randomized trial evaluating two treatment strategies for patients (pts) with aggressive lymphoma relapsing after or with progressive disease following primary therapy. The first randomization demonstrated that gemcitabine, dexamethasone, cisplatin (GDP) was non-inferior to and significantly less toxic than dexamethasone, cytarabine, cisplatin (DHAP) as salvage therapy (ASH 2012). Here we report the results of the second randomization, testing the ability of the CD20 antibody rituximab (R) administered post-ASCT to improve event-free survival (EFS) compared to no further therapy. Methods Pts with CD20-positive aggressive lymphoma who underwent ASCT after GDP or DHAP who had recovered from ASCT-related toxicities and who remained clinically progression-free within 3-5 weeks post-transplant were stratified by centre, salvage regimen received, response to salvage therapy (CR vs PR/SD) and prior treatment with R, and were randomized using a minimization algorithm to receive R 375 mg/m2every 2 months for 6 doses, or observation. Response assessment by CT scanning was required at 3,7,13 and 25 mos post-ASCT or as needed to evaluate possible disease recurrence. The primary endpoint of the second randomization was 2 year EFS; to detect an improvement by 15% (from 50 to 65%, hazard ratio 0.62) required 142 events, with a 2-sided α 0.05 and power 0.80. Because of the low event rate over the last year and a projected time of many years to reach the protocol specified event rate, the Data Safety Monitoring Committee approved a request for study closure and analysis, with 118 events recorded. Results 230 patients were randomized to R maintenance (115) or observation (115). Baseline patients and disease characteristics were well balanced between treatment arms: median age was 53 yrs, 28% were age >65 and 40% were female; 52% received GDP and 48% DHAP; 55% had an IPI score of 2 or more at relapse/progression, and 17% had transformed (TR) from previous indolent lymphoma. Response to salvage pre-ASCT: CR 24%, PR 58%; 70% had received R with chemotherapy prior to study enrolment, and 69% received R with protocol salvage treatment. All analyses are by intention to treat (ITT). To date, there have been 118 EFS events (R 53, observation 65). After a median follow-up of 63 months, 2 year EFS was 64% for pts treated with maintenance R vs 51% for those on observation (HR 0.74, 95% CI 0.48-1.14, p= 0.11); there was no difference in overall survival (OS) at 4 years (R 69%, observation 68%, p=0.64). Grade 3-4 neutropenia was reported in 36% of pts on R maintenance vs 25% during observation; and Gr 3-4 thrombocytopenia in 11% and 16%, respectively. Febrile neutropenia occurred in 10 pts (9%) on R maintenance and 2 pts (2%) on observation. Two year EFS was similar in subsets defined by stratification variables at randomization: GDP salvage therapy: R 57.0% vs observation 45.9% (HR 0.84, 95% CI 0.52-1.36), DHAP: R 70.0% vs observation 57.1% (HR 0.68, 0.39-1.18); response to salvage CR/CRu: R 69.0% vs 52.9% (HR 0.71, 0.32-1.6), PR: R 64.5% vs observation 57.4% (HR 0.90, 0.55-1.46); R use with prior treatment: R 53.7% vs observation 42.0% (HR 0.81, 0.54-1.22); no prior R: R 83.3% vs observation 70.6% (HR 0.64, 0.29-1.38). In multivariable analysis, only age >60 was significantly associated with EFS; ECOG performance status, treatment arm, stage and extra-nodal disease were not significant. Conclusion This evaluation of rituximab maintenance treatment every 2 mos for one year after ASCT for aggressive lymphoma failed to meet the study endpoint of improved EFS, compared to observation. Disclosures: Crump: Roche: Honoraria; Jansen-Ortho: Honoraria; Celgene: Honoraria; Lundbeck: Honoraria; Novartis: Research Funding; Seattle Genetics: Honoraria. Off Label Use: rituximab for maintenance therapy post autolgous transplant for lymphoma. Kuruvilla:Roche Canada Seattle Genetics, Janssen, Celgene, Lundbeck, Karyopharm: Honoraria. Kouroukis:Roche: Honoraria. Federico:MedImmune: Research Funding. Meyer:Lilly: Consultancy; Celgene: Consultancy.

Abstract 3681 Introduction: The First-Line Indolent Trial (FIT) demonstrated that administration of a single infusion of 90Y-RIT to follicular lymphoma grade 1 or 2 patients with CR or PR after induction chemotherapy could significantly prolong time to progression versus no further therapy. [JCO 2008;26(32):5156–63]. Recently, Fowler et al (ASH 2011, abstract 99) have reported on a chemoimmunotherapy approach followed by both RIT consolidation and rituximab maintenance for advanced stage FL patients with FLIPI risk factors >2. However, molecular status (PCR for t[14;18]) was not reported beyond one year of follow-up. We report a phase II study of safety and efficacy of 90Y-RIT following R-CHOP chemotherapy in patients with FLIPI=2–5, advanced stage FL. Maintenance rituximab is given every 3 months (m) post 90Y-RIT, for 24 m. Planned accrual is 33 evaluable patients with extensive two year molecular and immunologic follow-up. Novel insights into the biologic and immunologic effects of this combination regimen are presented. The primary endpoint of our study is the final complete response (CR) rate, defined according to the Cheson criteria and measured 3 m after day 1 of the 90Y-RIT therapy. Secondary outcomes included molecular remission and immunologic effects. Methods: In patients who had PCR detectable t[14;18] in baseline diagnostic specimens, quantitative real-time PCR was performed on blood and available bone marrow, at baseline, post 6xR-CHOP and Q 3 months post 90Y-RIT treatment for 24 m. The sensitivity of our PCR assay was 1 in 105cells. Flow cytometry for % B cell clonality was performed at the same time points. T and B cell counts, Ig levels and vaccine serology have been recorded pre and post treatment. Results: We have enrolled 26 patients with a median age of 54 yrs, 80% stage 4, 58%, intermediate FLIPI=2, 42% high FLIPI=3–5. Sixteen of 26 [62%] patients had a complete response (CR/CRu) to R-CHOP and the remaining 10 [40%] showed partial response (PR). One patient died due to sepsis prior to 90Y-RIT. Five patients with a partial response post R-CHOP converted to CRu post 90Y-RIT. A total of 19 of 24 (79%) patients who received 90Y-RIT, achieved CR/Cru. Post 90Y-RIT, three patients have relapsed. One other developed a secondary malignancy by 9m. The treatment has been most favourable for patients with FLIPI=2, where 13/14 (93%) remain progression-free [median follow-up= 32 m, range= 12.5–62 m]. There were no SAEs attributable to the 90Y-RIT treatment. Seventeen patients had PCR detectable t(14:18) translocations. Quantitative PCR measurements were concordant with flow cytometry. Of these, 16, were evaluated post 90Y-RIT and 15/16 (94%) of these patients became PCR negative in blood. Post 90Y-RIT, 2 patients showed increase in PCR levels and relapsed clinically. All remaining pts with PCR markers are PCR negative in blood as far as 24 m post 90Y-RIT. CD3+T cell counts remained normal, but CD19+B cells fell below the 1% detection level by flow cytometry during the two yrs of maintenance therapy post 90Y-RIT. Interestingly, mean IgG levels remained close to normal, but mean IgM levels fell below normal. Memory immune responses to measles and mumps were maintained post chemo-radiotherapy. Antibody titres to Rubella did not change significantly post 90Y-RIT. No HAMA response has been detected in any of the patients. Conclusions: We found effective eradication of follicular lymphoma from the blood and bone marrow of the high risk lymphoma pts with 2 or more FLIPI risk factors with first line treatment of 6xRCHOP and all but one of our evaluable patients (94%) achieved molecular remission in blood post 90Y-RIT. Molecular remission was sustained after 90Y-RIT up to 2 years, considerably longer than that reported by Fowler et al (ASH 2011, abstract 99). Longer follow-up with annual monitoring is planned to determine the precise response duration. The progression-free survival rates are similar or more favourable to previous reports [Blood 2006;108:1504–1508, J Clin Exp Hem. 2012; 52,no. 1]. IgG levels remain close to normal indicating that memory B cells are intact and this was consistent with no significant change in titres to common previously vaccinated pathogens such as rubella. The significance of persistent reductions on IgM levels is unclear. Acknowledgments: This study was sponsored by Bayer Canada and Spectrum Pharmaceuticals. Disclosures: Berinstein: Spectrum Pharma: Clinical Advisory Board Other. Off Label Use: Zevalin for first line treatment of aggressive follicular lymphoma.

Abstract 4981 Follicular lymphoma (FL) is an indolent lymphoma characterized by slow growth, initial response to treatment with inevitable relapse. Recent trials have demonstrated improved progression free survival (PFS) with rituximab-containing induction, maintenance rituximab (MR) or both. However, a small subset of patients (pts) will show disease progression during the induction or maintenance phase. We evaluated rituximab resistance (RR) including incidence, predictors, rate of transformation and prognosis for pts demonstrating RR. We defined RR as disease progression during induction, maintenance or within 6 months of last dose of rituximab. We retrospectively identified pts who received rituximab for symptomatic FL from July 2006 to April 2010 at 2 regional cancer centres. Those with a diagnosis of FL grades 1, 2 or 3a with first exposure to rituximab (induction, maintenance or both) were included. Exclusion criteria included FL grade 3b and previous rituximab exposure. Progression was ascertained from clinical notes or radiological investigation as per IWG Criteria (1999). Transformation was defined by confirmatory biopsy and clinical suspicion of transformation was also recorded (nodal growth, rapid rise in LDH or new B symptoms). PFS and overall survival (OS) were measured from initiation of rituximab induction. Log-rank statistics were used to identify univariate predictors for RR and Cox regression for multivariate analysis with outcome as time to early progression (RR). Of the pts screened, 132 met inclusion criteria (112 pts receiving primary therapy, 20 for ≥ 2nd line therapy); 22 pts (16.7%) demonstrated RR. Incidence of RR was similar for pts receiving primary therapy (17.0%). Pt characteristics for rituximab sensitive (RS) and RR groups are shown in Table 1. From induction, median follow-up was 33 months (range 9 to 61 months). In univariate analysis, high risk FLIPI score at induction was predictive of RR (p = 0.002). Partial response to induction (p = 0.082) trended to significance. Other factors not predictive of RR included: age, gender, high grade histology (grade 3a), previous chemotherapy received, time from diagnosis to induction therapy and anthracycline-based induction. A Cox regression model was constructed with FLIPI score and anthracycline-based induction as covariates. FLIPI score was independently and significantly predictive of RR (HR 2.43; 95% CI, 1.4 to 4.1; p = 0.001). Of the 22 pts who were RR, 18 (81.8%) required subsequent chemotherapy with only 9 achieving at least a partial response. Ten pts (45.5%) required more than 1 line of chemotherapy post-progression with a median time to next line of therapy of 9 months (95% CI, 2.9 to 15.1; see figure). Among the RR pts, 5 (22.7%) showed evidence of transformation on biopsy at initial resistance and an additional 3 (13.6%) were clinically suspected. Within 12 months of resistance, a further 3 pts in the RR group demonstrated biopsy-proven transformation. Overall, 10 pts (45.5%) transformed (biopsy-proven) with no transformation in the RS group. During follow-up, 9 deaths occurred, all in those with RR, 5 following biopsy-proven transformation. The median PFS and OS in the RR group were 17 months and 47 months, respectively. Table 1 Characteristics RS (n = 110) RR (n = 22) Age (MEAN) 58.9 58.9 Gender (females) 43.6% 40.9% High histological grade (3a) 8.4% 9.1% Previous treatment (yes) 15.5% 13.6% Alkylator 11.8% 13.6% Number of lines of previous treatment >1 4.5% 4.5% Mean time from diagnosis to induction (months) 30.3 28.1 Induction chemo R-CVP based 78.2% 86.4% R-CHOP based 18.2% 9.1% Other 3.6% 4.5% Response to induction PR 77.3% 72.7% CRu 12.7% 0% CR 10.0% 4.5% SD/PD 0% 22.7% FLIPI score at induction Low (0, 1, 2) 57.8% 18.2% Intermediate (3) 26.6% 40.9% High (4 or 5) 15.6% 40.9% Stage 3 or 4 88.2% 100% High tumor burden at diagnosis (GELF criteria) 43.6% 40.9% We demonstrated that the incidence of rituximab resistance in FL on first exposure to rituximab in our population was 16.7%. FLIPI score was predictive for RR and was independent of anthracycline-based induction regimens. Pts with RR had a high rate of histologic transformation (36.3% within 12 months post RR) and a shorter PFS/OS with a poor response to next therapy. Biopsies to confirm transformation should be performed in all pts demonstrating RR, and new therapies are needed for this group of pts. Disclosures: Crump: Ortho Johnson & Johnson: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees; Millennium Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees. Kukreti:Celgene: Honoraria; Ortho Biotech: Honoraria; Roche: Honoraria.

Abstract 2701 Introduction: The First-Line Indolent Trial (FIT) demonstrated that administration of a single infusion of 90Y-RIT to follicular lymphoma grade 1 or 2 patients with CR or PR after induction chemotherapy could significantly prolong remissions over no further therapy. Only 13% of patients in that trial received rituximab with their first-line chemotherapy and maintenance rituximab was not administered [JCO 2008;26(32):5156–63]. We report a phase II study of safety and efficacy of 90Y-RIT following R-CHOP chemotherapy in patients with high-risk advanced stage FL. Maintenance rituximab is given every 3 months post 90Y-RIT, for 24 months. The protocol accrues up to 33 evaluable patients. First-time novel insights into the biologic and immunologic effects of this combination regimen are presented. The primary endpoint of our study is the final complete response (CR) rate, defined according to the Cheson criteria and measured 3 months after day 1 of the 90Y-RIT therapy. The CR rate among similar risk advanced stage FL patients after R-CHOP is known to be about 20% [Blood 2005;106(12):3725–32]. The current study is designed to detect a significantly greater response rate of 40% with a p value of 0.05 and a power of 80%. Secondary outcomes included molecular remission and immunologic effects. Since molecular remission is associated with prolonged survival [Blood 105(9): 3428–3433], we analyzed quantitative PCR for up to two years post 90Y-RIT therapy on a subset of patients with baseline PCR detectable t(14:18) lymphoma. Methods: In patients who had PCR detectable t(14;18), quantitative real-time PCR was performed on blood and available bone marrow, at baseline, post 6xR-CHOP and Q 3 months post 90Y-RIT treatment for 24 months. The sensitivity of our PCR assay was 1 in 105 cells. Flow cytometry for % B cell clonality was performed at the same time points. T and B cell counts, Ig levels and vaccine serology have been recorded pre and post treatment. Genotyping of FCGR3A polymorphism was performed. Results: 24 pts have been enrolled with a median age of 54 yrs, 79% stage 4, 63% high FLIPI, 38%, intermediate FLIPI. 10 of 19 [53%] patients had a complete response (CR/CRu) to R-CHOP and the remaining 9 showed partial response (PR). One patient died due to sepsis prior to 90Y-RIT. 5 Patients with a partial response post R-CHOP converted to CRu post 90Y-RIT resulting in 14 of 18 (77%) patients who achieved CR/Cru. One patient progressed 6 m post 90Y-RIT, another relapsed at 12 months. One pt developed a secondary malignancy by 9 m. There were no SAEs attributable to the 90Y-RIT treatment. 16 of 24 (67%) had PCR detectable t(14:18) translocations. Quantitative PCR measurements were concordant with flow cytometry. 18 have been evaluated post 90Y-RIT. 12/13 (92%) of these pts became PCR negative in blood. Post 90Y-RIT, one pt relapsed and showed increase in PCR levels. All remaining pts with PCR markers are PCR negative in blood as far as 24 m post 90Y-RIT. CD3+T cell counts remained normal, but CD19+B cells fell below the 1% detection level by flow cytometry during the two yrs of maintenance therapy post 90Y-RIT. 16 patients have been followed for more than 6 m and 8 pts for 18–24 m with no significant increases in the CD19+ B cell numbers. Interestingly, mean IgG levels remained close to normal, but mean IgM levels fell below normal. Memory immune responses to measles and mumps were maintained post chemo-radiotherapy. Antibody titres to Rubella did not change significantly post 90Y-RIT. Conclusions: We found effective eradication of follicular lymphoma from the blood and bone marrow of the high risk lymphoma pts with first line treatment of 6xRCHOP and all but one of our evaluable patients (92 %) achieved molecular remission in blood post 90Y-RIT. Molecular remission was sustained with maintenance rituximab after 90Y-RIT up to 2 years. Longer follow-up will be necessary to determine the precise response duration. Our molecular response results compare favourably with those of the FIT trial. [JCO 2009;27(34):6094–00]. In our trial, IgG levels remain close to normal indicating that memory B cells are intact and this was consistent with no significant change in titres to common previously vaccinated pathogens such as rubella. A reduction in IgM levels indicating potential functional consequences in mounting primary humoral responses was documented. Disclosures: Off Label Use: Zevalin in first line follicular lymphoma.

Abstract 1357 Introduction: High dose chemotherapy (HDT) with autologous stem cell transplantation (ASCT) is historically effective treatment for patients with relapsed follicular lymphoma (FL) but relapse is common possibly due to contaminated stem cell grafts or inadequate eradication of primary disease. The addition of immunotherapy to HDT/ASCT may augment “in vivo” graft purging and achieve more effective eradication of minimal residual disease (MRD) post ASCT. This may lead to improved PFS and OS. Methods: We conducted 3 sequential prospective phase 2 trials of HDT/ASCT combined with immunotherapy in patients with relapsed FL from 1997–2010. Protocol 1 (n=13) used a-interferon 3 MU/m2 SC TIW for 2 years post ASCT. Protocol 2 (n=23) used a single infusion of rituximab (R) 375 mg/m2 for ‘in vivo’ purging 3–5 days pre-stem cell collection and 2 sets of 4 weekly R at 2 and 6 months post ASCT respectively. Protocol 3 (n=32) used 3 infusions of R 375 mg/m2 pre stem cell collection, followed by a-interferon 3 MU/m2 SC TIW × 2 years + R 375mg/m2 × 6 weekly infusions post ASCT. Eligibility included adult patients age 18–65 with stage III or IV follicular lymphoma grades 1–3a in first or second relapse. Salvage chemotherapy was either CHOP or DHAP depending on prior anthracycline exposure. High dose therapy was cyclophosphamide, carmustine, and etoposide (CBV). Minimal residual disease (MRD) assessment of t(14; 18) by quantitative PCR was serially performed in blood, PBSC and marrow. Studies were REB approved and all patients gave informed consent. Results: Sixty-eight patients from the 3 trials are included. Median age at study enrolment was 47 (30-71) and patients were a median of 31 mo (9-197) from diagnosis. 54% were male. Median FLIPI score was 2. Median # of prior chemotherapies was 1. Median number of cycles of salvage chemotherapy was 4 (2 - 10). 63 patients proceeded to ASCT. 5 patients were not transplanted either do to disease progression (2), or failed stem cell mobilization (3). Baseline characteristics of the patients enrolled in the three trials were similar, with the exception of rituximab(R) exposure in 22% of the patients in Protocol 3. 4 (6%) patients were lost to follow up. Median time to death or last follow was 84.5 mo (7-166). 32 patients (47%) relapsed and 20 (29%) have died. Median follow up times for each trial were 116, 99 and 57 mo. Actuarial median OS for Protocol 1 was 136 mo from enrolment and has not yet been reached for Protocols 2 and 3. Actuarial median PFS for Protocols 1 and 3 are 49 and 78 mo respectively and has not yet been reached for Protocol 2. MRD assessment by PCR was collected on 48 patients. Compared with study 1, the increased use of R pre SC collection improved in vivo purging by 2 logs. In Protocols 1 and 2, most still had detectable disease in the graft to a sensitivity of 1 cell/100,000. In Protocol 3, 56% had MRD negative apheresis products. 91% of 46 evaluable patients achieved molecular remission post stem cell transplant. Thirteen patients (19%) developed secondary malignancies post ASCT with a median time to malignancy of 88.5 mo (39-144). 1 patient developed 2 distinct malignancies. Secondary malignancies present were: MDS (4), AML (1), ALL (1), basal cell carcinoma (2), squamous cell carcinoma (3), thyroid cancer (1), non small cell lung cancer (1) and adenocarcinoma of unknown origin (1). Five (7%) patients transformed to DLBCL, with a median time to transformation post ASCT of 43.5 mo (17-142). The majority of patients who received rituximab immunotherapy pre and post ASCT developed persistent hypogammaglobulinemia. 17 patients (26%) developed interstitial pneumonitis or >2 respiratory complications post rituximab. 4 patients (6%) require monthly IVIG to treat recurrent respiratory infections. 10 patients (15%) developed herpes zoster 15% and 30% of patients in Protocols 1 and 3 did not complete the 2 years of alpha-IFN post ASCT secondary to one or more of depression, fatigue or cytopenias. Conclusion: HDT + ASCT combined with R +/− a-IFN produces durable PFS and molecular remissions but is associated with prolonged hypogammaglobulinemia and higher rates of interstitial pneumonitis. Three infusions of R pre SC collection achieve higher rates of molecular remission. Compliance with 2 years of a-IFN added to R post ASCT is poor and may not add clinical benefits to R alone. The additional role of HDT/ASCT in the era of R-chemotherapy for FL needs to be explored. Disclosures: No relevant conflicts of interest to declare.

Abstract 4770 BACKGROUND Relapse in the central nervous system (CNS) following initial treatment of diffuse large B-cell lymphoma (DLBCL) is an uncommon but fatal complication. However, the addition of rituximab improves the clinical outcome dramatically in DLBCL patients; its influence on CNS relapse is unproven. Aim This single centre retrospective study was conducted to investigate the incidence of CNS relapse, and to evaluate the impact of adding rituximab to standard CHOP (RCHOP) regimen without CNS prophylaxis in patients at risk of CNS relapse. PATIENTS AND METHODS All patients with DLBCL diagnosed from April 2002 to December 2007 at sunnybrook cancer center were retrospectively identified in the Cancer Database. Patients were included if they were >16 years old, had advanced stage (stage III /IV, or stage I /II with B symptoms, elevated LDH or bulky disease, were treated with RCHOP regimen with curative intent and were free of CNS involvement at diagnosis. CNS relapse was diagnosed by CSF cytology, radiology or clinically. Results A total of 155 patients were newly diagnosed with DLBCL and treated with RCHOP only. 22 pts were excluded, 20 had CNS prophylaxis and 2 pts had CNS involvement. 133 pts were eligible (69 male and 64 female) Median age was 64 (Age'60 was 59.4%). Stage III/IV was 69.9%. LDH was elevated in 59.4%. Bone marrow (BM) involvement and Extra nodal “>2 were 18.05% and 25.6% respectively. EN sites were: (liver 4.5%, Bone 6.8%, Pulmonary 4.5%, kidney 3.01%, cardiac 1.5%, intestine 2.3%, testicular 1.5%). The International Prognostic Index was high-intermediate/high in 55.6%. Pathologically transformed was 12.03% and were transformed from indolent histologies. BCL2 was positive in 65.4%, BCL6 was 48.9%, CD10 was positive in 49.6%, Ki-67 was >80% in 25%. All patients received RCHOP (Median 6 cycles, (range 2-8). Overall response (ORR) was 88.6%, CR/CRU 72.7% with a median follow up 24.6 months (range 2.6-75.5). 28 patients (21.05%) relapsed systemically. Two patients (1.5%) had a CNS relapse 1 brain parenchyma and 1 leptomeningeal one month after systemic relapse. The median time to CNS relapse was 10.4 mos (6.24-14.5 mos). In univariate risk factor analysis (LDH (p=0.8), IPI>3 (p=0.9), No of EN (p=0.9). Actuarial 5 y Overall Survival (OS) was 67.3% (95% CI (57-77%) and progression free Survival (PFS) was 65.7% (95% CI (52.3-78.6%). Conclusion Our data suggest that the addition of rituximab may reduce the risk of CNS relapse for poor risk patients likely through systemic control. Future prospective studies of rituximab-containing chemotherapies with CNS prophylaxis are warranted Disclosures: No relevant conflicts of interest to declare.

High-dose therapy and autologous stem cell transplantation (HDT/ASCT) and rituximab immunotherapy have been increasingly applied in the management of non-Hodgkin’s lymphomas (NHL). Although both approaches have been individually associated with B-cell depletion and hypogammaglobulinemia, the incidence, time course, and predictors of prolonged deficiencies following a combined treatment approach is unknown. Methods: We completed a series of prospective phase II studies of HDT/ASCT combined with rituximab for patients with relapsed follicular lymphoma (FL) or diffuse large B cell lymphoma (DLBCL). In two phase II trials of patients with FL (R/Tx and R-IFN/Tx), patients received 9 infusions of rituximab 375mg/m2 as both an in vivo purge and as maintenance post HDT/ASCT. In a trial with relapsed DLBCL or transformed lymphoma, patients received 8 infusions with rituximab 375mg/m2 only as part of the salvage chemotherapy regimen (R-ESHAP/Tx). Immunoglobulin levels were expressed as percentages with 100% representing the lower limit of normal at the institutional lab. Hypogammaglobulinemia was defined as Results: Fifty-one patients with FL were transplanted and were subsequently treated with maintenance rituximab (R/Tx study) or with maintenance rituximab (MR) and interferon-alpha (R-IFN/Tx). Twenty-six patients with relapsed aggressive-histology lymphoma were transplanted with the R-ESHAP salvage regimen. The mean age of the FL patients was 45 and the mean age of the aggressive-histology lymphoma patients was 49. The median number of prior therapies was 1 (range 1–3) for the FL patients and bone marrow involvement at study entry was present in 63%. The median number of prior therapies was 1 (range 1–5) for the aggressive-histology lymphoma patients and bone marrow involvement was present in 15%. Baseline IgG hypogammaglobulinemia was seen in 18% of patients with relapsed FL and 15% of patients with relapsed DLBCL/transformed lymphoma. Median baseline Ig levels in each study are displayed in Figure 1. Patients with FL undergoing HDT/ASCT and MR (combined R/Tx and R-IFN/Tx studies) had prolonged IgG hypogammaglobulinemia compared to patients with aggressive-histology lymphoma (log-rank p=0.0047; see Figure 2). The median time-to-recovery from hypogammaglobulinemia had not been reached after a median follow-up of 42 months. Similarly, IgM level recovery was delayed in the FL group compared to the aggressive-histology patients (p=.0001), although late recovery was noted for this Ig subtype (median time-to-recovery 36 months post-ASCT). Univariate analyses revealed that persistent hypogammaglobulinemia (at 24 months) was associated with FL histology (vs. DLBCL/transformed; p=0.006), bone marrow involvement at study entry (p=0.008), and hypogammaglobulinemia at study entry (p=0.026). Factors not associated with persistent hypogammaglobulinemia included: number of prior therapies, age, and administration of maintenance interferon-alpha. Post-transplant (>3 months) infections included herpes zoster reactivation (n=7) and pneumonia (n=8). One individual in the R-ESHAP/Tx study died of PCP pneumonia 4 months post HDT/ASCT. A relationship between grade III-IV infections and prolonged hypogammaglobulinemia was not evident on univariate analysis (data not shown). Conclusions: Patients with follicular lymphoma undergoing high dose therapy and stem cell transplantation together with rituximab maintenance are likely to experience a prolonged hypogammaglobulinemia whereas this is less likely with patients with aggressive-histology lymphoma undergoing similar doses of rituximab as part of their salvage therapy. Further research is required to elucidate the relative contributions of disease histology, bone marrow involvement, baseline hypogammaglobulinemia, timing of rituximab infusions or other factors not yet identified. Figure Figure Figure Figure

High-dose therapy and autologous stem cell transplantation (HDT/ASCT) are associated with prolonged remissions in relapsed follicular lymphoma (FL). Maintenance rituximab (R) has an accepted role in extending remissions and preclinical evidence suggests that its effects may be enhanced by the immunomodulatory capacity of interferon alpha (IFN-α). We designed a phase II study to incorporate in-vivo purging (with R) and post-transplant maintenance immunotherapy (with combined R and IFN-α) to prolong the remissions attained by HDT/ASCT. We present a planned analysis of 30 patients age ≤65 with 1–2 relapses of FL. Individuals received salvage CHOP or DHAP and proceeded with stem cell mobilization and HDT/ASCT with cyclophosphamide/carmustine/etoposide if they achieved ≥75% reduction in bulk and Results: Twenty-nine patients are assessable with a median follow-up for survivors of 3.1 years. Median age was 46 years (30–65) and median number of prior regimens was 2. Median response duration to prior therapy was 0.6 years and patients were enrolled 2.3 years (median) following diagnosis. The overall response rate to salvage was 93% (95% CI 83.7% to 100%) and n=23 proceeded to HDT/ASCT. Six patients did not undergo ASCT due to inadequate response (n=2), failed mobilization (n=2), cardiomyopathy (n=1), and patient withdrawal (n=1). Significant non-hematologic ASCT toxicities (grade 3/4) included: 5 episodes of interstitial pneumonitis, retinal vein occlusion (n=1), and thrombotic thrombocytopenic purpura (n=1). One patient was diagnosed with ALL 30 months following ASCT. Twenty-one individuals initiated IFN-α with 14 able to complete 2-years of maintenance (median dose of 3 × 106 units/m2 tiw). The most common reason for discontinuation was depression. At baseline, 19 patients had detectable markers by PCR. Despite in-vivo purging with R, 8 of 17 stem cell grafts had molecular disease, although graft contamination did not affect subsequent MR (p=NS). Of 16 assessable patients post-transplantation, 11/16 achieved MR prior to immunotherapy and 16/16 achieved MR during maintenance. Three patients have had a molecular relapse at 12, 18, and 36 months post-ASCT, with molecular preceding clinical relapse in 2/3 patients. Median progression-free survival for all patients is 50 months and median overall survival has not been reached. Conclusions: HDT/ASCT was feasible and well-tolerated in this high-risk population of relapsed FL. Molecular detection of lymphoma in the auto-graft did not preclude extended MR and clinical remissions post-transplant. This may be due to the enhanced clearance of minimal residual disease achieved by combination maintenance immunotherapy (R and interferon-α) post-ASCT. This is our third study in a sequential program of HDT/ASCT trials incorporating immunotherapy pre- and post-transplantation; we will present comparative evidence across the series to determine the added benefit of IFN-α in this latest approach.

Background: We previously reported that 20 patients with newly diagnosed, stage III/IV mantle cell lymphoma (MCL) treated prospectively with an in vivo rituximab purge, autologous stem-cell transplantation (ASCT) and rituximab maintenance experienced superior progression free survival (PFS) compared to 40 matched, historical controls treated with conventional chemotherapy. We now report extended follow-up from this trial. Methods: Eligible patients were treated with 4–6 cycles of CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) followed by high dose therapy (cyclophosphamide, carmustine and etoposide) and ASCT. Rituximab (375 mg/m2) was given once 5 days prior to stem cell collection (as an in vivo purge), and as two, 4-week maintenance courses 8 and 24 weeks after ASCT. Historical control patients matched for age, stage and gender were randomly selected from a lymphoma database maintained by the British Columbia Cancer Agency. Two control-patients were selected for every trial patient. All control patients were treated with either an anthracycline based-regimen, or a regimen containing fludarabine and cyclophosphamide. Results: Median follow-up is 5.3 years for the experimental cohort and 10.1 years for the control cohort. In the ASCT cohort, 8 patients have relapsed and 4 patients have died, 2 of progressive disease, 1 of a presumed cardiac event 7 months post-ASCT, and one of hepatitis B reactivation 11 months post-ASCT. Five-year OS and 5-year PFS were superior in MCL patients treated with ASCT+ rituximab compared to matched, historical MCL patients treated with conventional chemotherapy (5y OS 80% vs. 38%, P=0.0017; 5y PFS 72% vs. 19%, P=0.0001). One patient developed myelodysplastic syndrome 5.5 years after ASCT, and one patient was diagnosed with breast cancer 4.8 years after ASCT. Conclusions: With more than 5 years of follow-up MCL patients treated with ASCT plus rituximab continue to experience significantly improved OS and PFS compared to matched, historical controls. Overall Survival Overall Survival

High-dose therapy and autologous stem cell transplantation (HDT/ASCT) is associated with prolonged remissions in relapsed follicular lymphoma (FL). Molecular remission in the graft and post ASCT predicts durable remissions and is a desirable endpoint. Rituximab (R) as an in vivo purge prior to HDT/ASCT and as consolidation after ASCT may help achieve this. To study this question, patients with relapsed FL were enrolled in a prospective, non-comparative phase II study between January 1998 and April 2000. Methods: 23 consecutive patients age Results: Median cohort age at assessment was 50 yrs (32–57). Median number of prior regimens was 3 (1–7) and total treatment cycles was 9 (3–28). Median response duration to the preceding regimen was 10 months (1–86). Transplants were a median 2.4 years after diagnosis. At median follow-up of 4.5 yrs (1.3–6.3), there have been 10 relapses at a median of 2.8 yrs. 3 patients have died, 2 with relapse and one of presumptive sudden cardiac death. Significant toxicities were seen: 11 episodes of pneumonia (1 fungal), 4 episodes of herpes zoster (1 grade 3), 4 early episodes of grade 3/4 interstitial pneumonitis, and 1 episode each of grade 4 TTP and grade 3 optic neuritis. One patient developed AML at 4.7 years post-ASCT. Immune recovery has been delayed; at 600 days post ASCT, 16/18 (89%) of evaluable patients had not recovered IgG levels to normal. At baseline, 12/23 patients had detectable markers by PCR analysis (sensitivity 0.01%) for t(14;18) or patient-specific VDJ rearrangements in marrow or blood. Of these, all achieved at least a brief molecular remission in blood and marrow, 11/12 doing so pre-R consolidation. 5/12 patients have since relapsed at a median 3 yrs (2–4.5) post ASCT. Molecular relapse preceded clinical in 3/5 cases. 6/12 have had prolonged (median 4.8 yrs, range 3–5) molecular and clinical remissions. Median event-free survival for all patients is 63 months, with median overall survival not reached. Conclusions: HDT/ASCT with R for in vivo purging and post-ASCT consolidation for relapsed FL is feasible and associated with prolonged clinical and molecular remission. Delay in recovery of humoral immunity may play a role in the high incidence of infectious complications. A single infusion of Rituximab for in-vivo purging did not eradicate PCR detectable disease in the graft, although sustained molecular remissions of at least 24 months were achieved in 75% of evaluable patients post transplant, possibly due to post ASCT Rituximab consolidation. Whether this translates into survival benefit will require longer follow-up and further comparative studies.

High-dose therapy and autologous stem-cell transplantation (HDT/ASCT) is associated with long remissions in relapsed follicular lymphoma (FL). Molecular remission in the graft and post ASCT is predictive of durability of remission. Thus, a simple, reliable method of quantitation of minimal residual disease (MRD) would improve prognostication in these patients. Real-time polymerase chain reaction (PCR) can quantitatively detect FL markers in blood and BM with a reproducible sensitivity of 0.01%. Fluorescent probe systems are used to perform real-time PCR; however, these techniques are expensive and do not allow melt curve analysis of the PCR product. We describe here a less expensive SYBR green detection method which also enables us to check clonal identity, and yields sensitive, reliable quantitative results for monitoring MRD. We compared our real-time PCR results with previously determined conventional nested PCR results in a series of samples collected from patients enrolled in one of our clinical trials. Methods: Patients with relapsed FL (n=14) were enrolled in a prospective study of HDT/ASCT followed by maintenance Interferon-a 3x106 U/m2 SC 3 x weekly for 2 yrs. DNA extracts were collected at enrolment from samples of lymph node, BM and blood to determine detection of BCL2/JH translocation. Stem-cell graft DNA and serial post-ASCT blood and BM DNA samples were collected. BCL2/JH was qualitatively determined by nested PCR using standard primers and analyzed by gel electrophoresis. For patients in whom BCL2/JH was not detectable, clone-specific IgH region primers were used where possible. Quantitative PCR was performed using QuantiTect SYBR Green kits. Melt curve analysis was performed to confirm amplification specificity and clonal identity of each sample. All samples were analyzed in triplicate for accuracy. Results: At a median follow-up of 75 months (9–91), 10/14 patients are alive. Of these, 3 are free from progressive disease. Median progression-free survival is 42 months; median overall survival has not been reached. We were successful in defining molecular markers for 12/14 patients. In 11/12, we were able to PCR amplify the BCL2/JH translocation. In one patient, allele-specific clonal IgH was analyzed with patient specific oligonucleotides. Median BCL2/JH contamination of stem-cell grafts was 0.1% (range 0.01% – 13%). 6 patients who had measurable graft contamination became molecularly negative in blood and BM within 12 months after ASCT. Increasing fractions of BCL2/JH positive blood and BM cells reliably predicted morphologic and clinical relapse. Correspondingly, the 3 patients free of disease progression have sustained very low levels of molecular evidence of FL. In one illustrative case, a patient had a long-term low-level persistence of tumour cells in BM and blood with no clinical symptoms for 70 months post-ASCT. At 76 months there was a 10-fold increase in the quantity of PCR product in both BM and blood. Radiological evidence of relapse was observed 3 months later. The patient underwent spontaneous disease regression, which was paralleled by a 10-fold decrease in PCR product quantity at 82 months. Conclusions: Our simplified method of detection using quantitative PCR yields reliable measurements of graft contamination and MRD post ASCT, which may be useful in predicting patient relapse.

Background: Patients with relapsed or refractory aggressive B-cell lymphoma have a poor prognosis. If sensitive to salvage chemotherapy, high dose therapy followed by autologous stem cell transplant (ASCT) can result in long-term survival for some. Unfortunately, 50% of patients are insensitive to standard salvage regimens and thus are ineligible for ASCT. Hypothesis: Combining Rituximab with ESHAP may improve chemosensitivity and ASCT eligibility, of patients with CD20+, relapsed or refractory aggressive lymphoma, or CD20+ transformed indolent lymphoma. Objectives: The primary outcome was response rate (CR + CRu + PR) to R-ESHAP. Secondary outcomes were toxicity, ability to undergo ASCT, presence of minimal resdidual disease in the stem cell harvest, and progression free and overall survival. Methods: Eligible patients were 16–65 years old, had ECOG performance status 0–2, and had pathologically confirmed CD20+, relapsed/refractory aggressive lymphoma or transformed indolent lymphoma. Patients with refractory disease had a PR with initial anthracycline-based therapy. ESHAP (etoposide 40mg/m2 day 1–4, solumderol 500mg day 1–5, cytosine arabinoside 2g/m2 day 5, cisplatin 25mg/m2 day 1–4) was given every 28 days with GCSF support until < 15% bone marrow involvement was achieved (2–4 cycles). Eight infusions of Rituximab (375mg/m2) were administered weekly concurrent with the first 2 cycles of ESHAP. GCSF mobilized stems cells were collected on day 10 or 11 when < 15% bone marrow involvement was achieved (mainly in cycles 1 or 2). Results: Preliminary results of 14 of 44 planned patients are presented. Median age was 54 years (range 42–64). All pts had CD20+ aggressive lymphoma, 6 had relapsed aggressive lymphoma, 2 had refractory aggressive lymphoma (with PR after initial therapy) and 6 had transformed indolent lymphoma. Twelve of 14 patients were stage III/IV. The response rate to R-ESHAP was 93% (2 CR, 2 CRu, 9 PR, 1 PD). Thirteen of 14 patients proceeded to ASCT. Data on minimal residual disease in the stem cell harvest is pending. Grade 3–4 thrombocytopenia, neutropenia and anemia occurred with 53%, 31%, and 20% of R-ESHAP cycles respectively. Two of 14 patients had febrile neutropenia, one with bacteremia. Two patients had non-neutropenic bacteremia (1 with septic shock). There were no toxic deaths. Median follow-up post-ASCT is 5 months (range 1–18). Two patients progressed 4 and 5 months post-ASCT respectively. There were 3 deaths, 2 lymphoma related and 1 due to accelerated Parkinson’s Disease. Median PFS has not yet been reached. Conclusions: This trial is still accruing patients. However, preliminary results are promising. R-ESHAP appears to be well tolerated with a high response rate. This regimen may enable more patients with relapsed/refractory aggressive lymphoma or transformed indolent lymphoma to proceed to ASCT.